Unsaturate amino compounds for use as anticancer and antiprotozoic agent

ABSTRACT

The invention relates to compounds of formula (I), R 1  and R 2 , each independently of the other, are selected from lower alkyl that is unsubstituted or substituted by one or more fluorine atoms which are not linked to the carbon atom of R 1  or R 2  bonding the nitrogen; from lower alkenyl wherein the double bond does not originate from the carbon atom that is bonded to a nitrogen bonding R 1  or R 2  ; from lower alkynyl wherein the triple bond does not originate from the carbon atom that is bonded to a nitrogen bonding R 1  or R 2  ; from cycloalkyl; and from cycloalkyl-lower alkyl; with the proviso that not more than one of the two radicals R 1  and R 2  is methyl; or salts thereof. The mentioned compounds are pharmacologically active against disorders that are responsive to a recuction in intracellular polyamines, such as tumours or protozoal diseases.

This application is filed under 35 U.S.C. §371 as a national phaseapplication of PCT/EP94/04230, filed Dec. 14, 1994, which was based onSwitzerland Application No. 3876/93-9, filed Dec. 27, 1993.

The invention relates to N¹,N¹⁴ -disubstituted tetraazatetradec-7-enesand their salts, to processes for the preparation of those compounds, topharmaceutical compositions comprising those compounds, and to the useof those compounds in the therapeutic treatment of the human or animalbody or in the preparation of pharmaceutical compositions.

Polyamines, for example spermine, spermidine and analogues thereof, havefor some time been the subject of intensive investigation as regardstheir biological properties, especially as regards proliferativeprocesses. An early finding has been that higher levels of polyaminesare to be found in cells that are dividing, for example in cancer cells,than in cells that are stable.

Such phenomenological observations have led to the conclusion thatpolyamines are necessary for cell proliferation.

The concept of influencing the polyamine level in cells has thereforebeen made use of in chemotherapy, for example of cancerous diseases.

Surprisingly, it has now been found that the compounds of the presentinvention have especially valuable properties that can be usedpharmacologically.

The compounds according to the invention are compounds of formula I##STR2## wherein R₁ and R₂, each independently of the other, areselected from lower alkyl that is unsubstituted or substituted by one ormore fluorine atoms which are not linked to the carbon atom of R₁ or R₂bonding the nitrogen; from lower alkenyl wherein the double bond doesnot originate from the carbon atom that is bonded to a nitrogen bondingR₁ or R₂ ; from lower alkynyl wherein the triple bond does not originatefrom the carbon atom that is bonded to a nitrogen bonding R₁ or R₂ ;from cycloalkyl; and from cycloalkyl-lower alkyl; with the proviso thatnot more than one of the two radicals R₁ and R₂ is methyl; or saltsthereof.

Within the context of the present Application, the general terms usedhereinbefore and hereinafter have preferably the following meanings:

Lower alkyl has especially up to a maximum of 7 carbon atoms, isbranched or unbranched and is preferably methyl or especially C₂ -C₇alkyl, such as ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl,tert-butyl, n-pentyl, neopentyl, n-hexyl or n-heptyl, especially methylor more especially C₂ -C₄ alkyl, such as ethyl, n-propyl, isopropyl,n-butyl, sec-butyl or isobutyl, with ethyl and propyl, such as n-propyl,being especially preferred.

Those radicals are preferably unsubstituted but may also be substitutedby one or more, preferably up to three, fluorine atoms, for example asin 2,2,2-trifluoroethyl. The fluorine atom is not bonded to the carbonatom in R₁ or R₂ that is bonded to the nitrogen in formula I.

Lower alkenyl has especially from 3 to 7, preferably 3 or 4, carbonatoms and is, for example, allyl or crotyl.

Lower alkynyl has especially from 3 to 7, preferably 3 or 4, carbonatoms and is, for example, propyn-2-yl or 2-butyn-1-yl.

In lower alkenyl and lower alkynyl, an unsaturated bond must notoriginate from the carbon atom that is bonded to a nitrogen atom bondingR₁ or R₂, as unstable compounds are otherwise formed.

Cycloalkyl has preferably from 3 to 7, especially from 3 to 5, carbonatoms and is especially cyclopropyl or cyclobutyl.

Cycloalkyl-lower alkyl contains as cycloalkyl especially a radicalhaving from 3 to 5 carbon atoms, especially cyclopropyl or alsocyclobutyl, and as lower alkyl radical preferably a radical as definedabove, especially C₁ -C₃ alkyl, for example methyl, ethyl, n-propyl orisopropyl, more especially C₁ -C₂ alkyl. Preference is given to2-cyclopropylethyl, cyclobutyl-methyl or, especially, cyclopropylmethyl.

On account of their basic properties, salts of compounds of formula Iare especially acid addition salts, but may also be mixed salts.

Salts are especially the pharmaceutically acceptable, that is to saynon-toxic, salts of compounds of formula I, that is to say especiallythe corresponding acid addition salts with acid anions that are nottoxic (at the dose in question).

Such salts are formed, for example, by compounds of formula I withinorganic acids, for example hyclrohalic acids, such as hydrochloricacid or hydrobromic acid, sulfuric acid or phosphoric acid, or withorganic carboxylic, sulfonic, sulfo or phospho acids or N-substitutedsulfamic acids, for example acetic acid, propionic acid, glycolic acid,succinic acid, maleic acid, hydroxymaleic acid, methylmaleic acid,fumaric acid, malic acid, tartaric acid, gluconic acid, glucaric acid,glucuronic acid, citric acid, benzoic acid, cinnamic acid, mandelicacid, salicylic acid, 4-aminosalicylic acid, 2-phenoxybenzoic acid,2-acetoxybenzoic acid, embonic acid, nicotinic acid or isonicotinicacid, and also with amino acids, such as the 20 α-amino acids involvedin the synthesis of proteins in nature, for example glutamic acid oraspartic acid, and also with methanesulfonic acid, ethanesulfonic acid,2-hydroxyethanesulfonic acid, ethane-1,2-disulfonic acid,benzenesulfonic acid, 4-methylbenzenesulfonic acid,naphthalene-2-sulfonic acid, naphthalene-1,5-disulfonic acid, 2- or3-phosphoglycerate, glucose-6-phosphate, N-cyclohexylsulfamic acid (withthe formation of cyclamates), or with other acid organic compounds, suchas ascorbic acid. Carbonates or hydrogen carbonates are also possible.

The mixed salts include, for example, salts of compounds of formula Iwith di- or tri-valent acids that have acid radicals having differentdissociation constants, such as citric acid or phosphoric acid, where,for example, one or two protons of those acids have been replaced bycations, such as alkali metal cations, for example Na⁺ or K⁺, so thatthe corresponding salts still contain the corresponding cations as wellas the compound of formula I and the corresponding acid anions.

The terms "compounds" and "salts" expressly include also individualcompounds or individual salts.

For isolation or purification it is also possible to usepharmaceutically unacceptable salts, for example picrates orperchlorates. Only the pharmaceutically acceptable, non-toxic salts areused therapeutically, and those are therefore preferred.

The central double bond in the compounds of formula I is always in the E(=trans) form indicated. If asymmetric carbon atoms are present in oneor both of the radicals R₁ and R₂, then the corresponding compounds maybe in the form of isomeric mixtures, for example in the form ofdiastereoisomeric mixtures or racemates, or in pure form.

The compounds of the present invention have especially valuablepharmacological properties. In particular, it has surprisingly beenfound that the compounds of formula I effect a reduction in theintracellular concentrations (pool) of natural polyamines, such as,especially, putrescine, spermidine and spermine. This brings about aslowing down or suppression of cell divisions, especially a reduction inor cessation of the growth of growing (especially rapidly growing)tissues.

The reduction in the intracellular levels of polyamines, especially ofputrescine, spermidine and/or sperminc, is probably based primarily onthe fact that the compounds of formula I reduce the activities ofbiosynthetic enzymes of polyamine biosynthesis, ornithine decarboxylase(ODC) and/or S-adenosylmethionine decarboxylase (SAMDC). Additionally oralternatively, the compounds of formula I can bring about anacceleration in the metabolic decomposition of natural polyamines; forexample, induction of spermidine-spermine-acetyl-transferase, which inprinciple can even be super-induced, may be possible.

The reduction in the intracellular concentration of polyamines may bedemonstrated, for example, as follows (see C. W. Porter et at., CancerRes. 45, 2050-2057 (1985)):

Mouse ascites L1210 leukaemia cells are cultured (at 37° C.) inRPMI-1640 medium (which contains, per litre, 100 mg of Ca(NO₃)₂, 400 mgof KCl, 100 mg of MgSO₄.7H₂ O, 6000 mg of NaCl, 2000 mg of NaHCO₃, 801mg of Na₂ HPO₄, 242 mg of L-Arg.HCl, 50 mg of L-Ash, 20 mg of L-Asp, 50mg of L-Cys, 300 mg of L-Gln, 20 mg of L-Glu, 10 mg of Gly, 18.2 mg ofL-His.HCl.H₂ O, 20 mg of L-hydroxyproline, 50 mg of L-Leu, 40 mg ofL-Lys.HCl, 15 mg of L-Met, 15 mg of L-Phe, 20 mg of L-Pro, 30 mg ofL-Ser, 20 mg of L-Thr, 5 mg of L-Trp, 20 mg of L-Tyr, 20 mg of L-Vat, 1mg of 1-aminobenzoic acid, 0.2 mg of biotin, 3 mg of choline chloride, 1mg of folic acid, 35 mg of i-inositol, 1 mg of nicotinamide, 0.25 mg ofpantothenoic acid calcium salt, 1 mg of pyridoxine.HCl, 0.2 mg ofriboflavin, 1 mg of thiamine. HCl, 0.005 mg of vitamin B₁₂, 2000 mg ofglucose, 1 mg of glutathione, and 5 mg of phenol red), which alsocontains 2 % 4-(2-hydroxyethyl)-1-piperazineethanesulfonicacid/3-(N-morpholino)propanesulfonic acid, 1 mM aminoguanidine and 10%NuSerum (Collaborative Research Inc., Lexington, Mass.). The cells arecultured under a humid 5% carbon dioxide atmosphere at 37° C. either inglass culture tubes (2 ml) or in 25 or 75 cm² tissue culture bottles ina total volume of 15 or 50 ml, respectively. The cultures are treatedwith the compounds of formula I or with 0.1 mM spermidine (controls:without corresponding compounds) during the logarithmic growth phase(0.5 to 1×10⁵ cells/ml). The number of cells is measured by electronicparticle counting (Model ZF Coulter Counter, Coulter Electronics,Hialeah, Fla.) and confirmed from time to time by haemocytometermeasurements. The viability of the cells is measured by trypan blueexclusion measurement (0.5% in unbuffered 0.9% sodium chloride solution).

For polyamine determination, the cell samples are washed twice in coldPBS (="Phosphate Buffered Saline"--contains per litre 8000 mg of NaCl,200 mg of KCl, 1150 mg of Na₂ HPO₄.2H₂ O, 200 mg of KH₂ PO₄, 100 mg ofMgCl₂.H₂ O, 200 mg of MgSO₄.H₂ O and CaCl₂ ; pH 7.2), and an aliquot of10⁷ cells is removed for polyamine determination. The cells are formedinto a pellet, and the PBS supernatant is removed carefully using acotton wool swab. The pellet containing the cells is then maintainedtogether with 0.5 ml of 0.6M perchloric acid for 30 minutes at 4° C. andis then centrifuged for 3 minutes at 12,000 g using a microcentrifuge.The supernatant is frozen at -20° C. until the HPLC analysis. For thatpurpose, the polyamines in a 50 μl sample of the perchloric acid extractare separated over an HPLC system using a glass "Microbore Column"having a diameter of 2.8 mm, which is packed to a height of 2 cm withTLC-4A-cation exchange resin (Durrum Chemical Corp., Palo Alto, Calif.).The column temperature is maintained at 65° C. by means of a water bathwith circulating water. The column is eluted at a flow rate of 16 ml/hwith an initial column pressure of 34.45 bar, which decreases as theionic strength of the elution buffer increases. Buffer 1 (which contains0.2M boric acid, 0.5M NaCl, 0.03% Brij 35 (polyoxyethylene monolaurylether, the number of ethyleneoxy radicals is approximately 23; PierceChemical Co., Rockford, Ill.) and 0.0001% octanoic acid, pH adjusted to6.0 with saturated KOH) is flushed through the column for 4 minutes.Buffer 2 (which contains 0.2M boric acid, 2.15M NaCl, 0.03% Brij 35 and0.0001% octanoic acid, pH adjusted as above) is pumped through for 6minutes. Buffer 3 (which contains 0.2M boric acid, 2.9M NaCl and 0.0001%octanoic acid (pH adjusted as above)) is likewise applied for 6 minutes.The column is re-equilibrated for 10 minutes with buffer 1, before thenext sample is introduced. The column eluate is derivatised with 0.05%o-phthalaldehyde (Durrum Chemical Corp.) in 0.4M borate buffer (pH10.4)/1 mM 2-mercaptoethanol/0.09% Brij 35. The flow rate foro-phthalaldehyde is 8 ml/h. The derivatised eluate is examined for itspolyamine content by being passed through the flow cell of aflow-measuring device (Fluoro-Monitor; American Instrument Co., SilverSpring, Md.) with a fixed excitation wavelength of 360 nm and anemission wavelength of 570 nm. The data are determined using aHewlett-Packard Model 3385A automation system. The variance of thesystem for a standard with known concentrations of putrescine,spermidine and sperminc hydrochloride is lower than 5%. The sensitivityof the HPLC system is approximately 50 pmol/50 gl sample (10⁶ cells).

Using this method it is possible to observe a reduction in the polyaminelevels with the compounds of formula I of the present invention. Inparticular, at a concentration of a compound of formula I of from 5 to50 μM, for example 10 μM, the average levels of putrescine, spermidineand spermine are each reduced to less than 50% of the control values,especially to from 1 to 40% in the case of putrescine and spermidine andfrom 20 to 70% in the case of spermine.

As polyamine antimetabolites, the compounds of formula I haveantiproliferative properties which can be demonstrated, for example, bymeans of the inhibitory action on the growth of human T24 bladder cellcarcinomas. This is demonstrated by incubating the cells in "Eaglesminimal essential medium" (see Eagle, H., Science 130, c 1432-1437(1959)), to which 5% (v/v) foetal calf serum is added, in a humidifiedincubator at 37° C. and 5% by volume CO₂ in the air. The carcinoma cells(1000-1500) are transferred to 96-well microlitre plates and areincubated overnight under the said conditions. The test compound isadded in serial dilutions on day 1. The plates are incubated under thesaid conditions for 5 days. During that period, control cultures undergoat least 4 cell divisions. After incubation, the cells are fixed with3.3% (weight/volume=w/v) aqueous glutaraldehyde solution, washed withwater and stained with 0.05% (w/v) aqueous methylene blue solution.After washing, the dye is eluted with 3% (w/v) aqueous hydrochloricacid. The optical density (OD) per well, which is directly proportionalto the number of cells, is then measured by means of a photometer(Titertek multiskan) at 665 nm. The IC₅₀ values are calculated by meansof a computer system using the formula ##EQU1## The IC₅₀ value isdefined as the concentration of active ingredient at which the number ofcells per well at the end of the incubation period is only 50% of thenumber of cells in the control cultures.

For compounds of formula I, IC₅₀ values in the range of from 2×10⁻⁴ to5×10⁻⁸ M, especially in the range of from 3×10⁻⁵ to 10⁻⁷ M, areobtained.

The compounds of formula I are therefore especially suitable for the(therapeutic or preventive) treatment of pathological conditions thatare responsive to a reduction in the concentration of polyamines incells (intracellular polyamine concentration), for example proliferativedisorders, especially benign and malignant tumour disorders. They canbring about the regression of tumours and also prevent the spread oftumour cells (metastasisation) and the growth of micrometastases.Moreover, they can be used, for example, for treating protozoalinfections, such as, for example, trypanosomiasis, malaria, or pulmonaryinflammation caused by Pneumocystis carinii.

The compounds of formula I can be used as polyamine antimetaboliteseither on their own or in combination with other pharmacologicallyactive substances. They may be combined with, for example, (a)inhibitors of one or more enzymes of polyamine biosynthesis, for exampleornithine decarboxylase or S-adenosylmethionine decarboxylaseinhibitors, (b) inhibitors of protein kinase C, (c) inhibitors oftyrosine protein kinase, (d) cytokines, (e) negative growth regulators,(f) aromatase inhibitors, (g) antioestrogens or (h) conventionalcytostatic active ingredients.

Preference is given to compounds of formula I wherein R₁ and R₂, eachindependently of the other, am selected from C₂ -C₇ alkyl, especially C₂-C₄ alkyl, such as ethyl, n-propyl, isopropyl, isobutyl or n-butyl; C₃-C₇ alkenyl wherein the double bond does not originate from the carbonatom that is bonded to a nitrogen bonding R₁ or R₂, especially C₃ -C₄-alkenyl, such as allyl; C₃ -C₇ alkynyl wherein the triple bond does notoriginate from the carbon atom that is bonded to a nitrogen bonding R₁or R₂, especially C₃ -C₄ alkynyl, such as propargyl; and C₃ -C₅cycloalkyl-C₁ -C₂ alkyl, especially C₃ -C₄ cycloalkylmethyl, such ascyclopropylmethyl; as well as from C₃ -C₅ cycloalkyl, such ascyclopropyl or cyclobutyl; or salts thereof.

Greater preference is given to compounds of formula I wherein R₁ and R₂,each independently of the other, are selected from C₂ -C₄ alkyl, such asethyl, n-propyl, n-butyl or isobutyl, and C₃ -C₄ alkenyl wherein thedouble bond does not originate from the carbon atom that is bonded to anitrogen bonding R₁ or R₂, such as allyl or crotonyl, and also from C₃-C₄ alkynyl wherein the triple bond does not originate from the carbonatom that is bonded to a nitrogen bonding R₁ or R₂, such as propargyl,and (also) from C₃ -C₄ cycloalkylmethyl, such as cyclopropylmethyl,especially with the proviso that R₁ and R₂ together have not more than 6carbon atoms, or salts thereof.

Special preference is given to compounds of formula I wherein R₁ and R₂,each independently of the other, are selected from ethyl, n-propyl,isopropyl, n-butyl, isobutyl and allyl, or salts thereof, especiallythose compounds wherein R₁ and R₂ together have 4, 5 or 6 carbon atoms,or salts thereof.

Very special preference is given to compounds of formula I wherein R₁ isethyl and R₂ is selected from ethyl, n-propyl, n-butyl, isobutyl andallyl, or salts thereof.

Most especially preferred are the individual compounds of formula Imentioned in the Examples, especially those wherein R₁ and R₂ togetherhave from 4 to 6 carbon atoms, most especially those wherein neither R₁nor R₂ is lower alkynyl and wherein R₁ and R₂ are each bonded viamethylene (--CH₂ --), or salts thereof.

The compounds according to the invention are prepared for example by

a) nucleophilically substituting an amino compound of formula II##STR3## wherein R₂ is as defined for compounds of formula I and anyfunctional groups that are not to take pan in the reaction are, ifnecessary, in protected form, with a compound of formula III

    R.sub.1 --X                                                (III),

wherein R₁ is as defined for compounds of formula I and X is anucleofugal leaving group, with only one of the radicals R₁ and R₂ beingmethyl, and removing any protecting groups that are present, or

b) for the preparation of compounds of formula I wherein R₁ and R₂ arethe same and each have one of the meanings given in the definition ofthose radicals in compounds of formula I, other than methyl,nucleophilically substituting a diamine of formula IV ##STR4## whereinany functional groups that are not to take part in the reaction are, ifnecessary, in protected form, with a compound of formula V

    R.sup.X --X                                                (V),

wherein R^(X) has one of the meanings given for R₁ or R₂ in compounds offormula I, other than methyl, and X is a nucleofugal leaving group, andremoving any protecting groups that are present, or

c) for the preparation of compounds of formula I wherein R₁ and R₂ areeach independently of the other a radical R₁ or R₂ that is bonded via amethylene group (--CH₂ --) belonging to the radical in question,reacting a diamide of formula VI ##STR5## wherein R₁ ' and R₂ ' are eachindependently of the other the radicals, minus the above-mentionedmethylene group, which are complementary to the last-defined radicals R₁and R₂, and any functional groups that are not to take part in thereaction are, if necessary, in protected form, with selective reductionof the two amide groups, and, if necessary, removing any protectinggroups that are present, or

d) reacting a hexahydropyrimidine derivative of formula VII ##STR6##wherein Rx is one of the radicals indicated for R₁ and R₂ under formulaI and Y is a divalent protecting group, with an olefin of formula VIII##STR7## wherein Q and Q' are each independently of the other anucleofugal leaving group, with nucleophilic substitution of the twonucleofugal leaving groups, and removing any protecting groups that arepresent,

and, if desired, converting an obtainable free compound of formula Iinto its salt, convening an obtainable salt of a compound of formula Iinto the free compound or into a different salt of a compound of formulaI, and/or separating obtainable mixtures of isomers into the individualisomers.

In the following more detailed description of the preferred processes,R₁ and R₂ are as defined for compounds of formula I, unless indicatedotherwise.

Process a) (alkylation): In the starting materials of formula II, allthe nitrogen atoms are preferably in mono-protected form, so that thehydrogen that is to be replaced by R₁ is still present; in that case the4 nitrogen atoms are each bonded to a protecting group instead of to oneof the 4 hydrogen atoms shown in formula II that are not to be reacted.

The protecting groups are protecting groups that can be removed withoutreduction of the central double bond in the molecule of formula IItaking place.

The protecting groups for functional groups in starting materials thereaction of which is to be avoided, that is to say amino and iminogroups and hydroxy groups, include especially those protecting groups(conventional protecting groups) that are conventionally employed in thesynthesis of peptide compounds and also of cephalosporins andpenicillins as well as nucleic acid derivatives. Those protecting groupsmay already be present in the precursors and are intended to protect thefunctional groups in question from undesired secondary reactions, suchas acylations, esterifications, oxidations, solvolyses, etc. Acharacteristic of protecting groups is that they are readily removable,that is to say without undesired secondary reactions taking place, forexample by solvolysis, by reduction (without the simultaneous reductionof double bonds or of triple bonds that may be present), by photolysisor enzymatically, for example also under physiological conditions. Acharacteristic of protecting groups is that they are not present in theend products.

The protection of functional groups by such protecting groups, theprotecting groups themselves and the reactions for their removal aredescribed, for example, in standard works, such as J. F. W. McOmie,"Protective Groups in Organic Chemistry", Plenum Press, London and NewYork 1973, in Th. W. Greene, "Protective Groups in Organic Synthesis",Wiley, New York 1981, in "The Peptides"; Vol. 3 (E. Gross and J.Meienhofer, eds.), Academic Press, London and New York 1981, in"Methoden der organischen Chemie", Houben-Weyl, 4th edition, Vol. 15/I,Georg Thieme Verlag, Stuttgart 1974, and in H.-D. Jakubke and H.Jescheit, "Aminosauren, Peptide, Proteine", Verlag Chemie, Weinheim,Deerfield Beach and Basle 1982.

A protected amino or imino group is protected by a monovalentamino-protecting group, for example in the form of an acylamino,arylmethylamino, 2-acyl-lower alk-1-enylamino or silylamino group.Divalent protecting groups bridging two adjacent nitrogen atoms are alsopossible.

Hereinafter an "amino"-protecting group is always to be understood asmeaning also a corresponding imino-protecting group.

In an acylamino group acyl is, for example, the acyl radical of anorganic carboxylic acid having, for example, up to 18 carbon atoms,especially of a lower alkanecarboxylic acid that is unsubstituted orsubstituted by, for example, halogen or aryl, or of a benzoic acid thatis unsubstituted or substituted by, for example, halogen, lower alkoxyor nitro, or preferably of a carbonic acid semiester. Such acyl groupsare preferably lower alkanoyl, such as formyl, acetyl, propionyl orpivaloyl, halo-lower alkanoyl, for example 2-haloacetyl, such as2-chloro-, 2-bromo-, 2-iodo-, 2,2,2-trifluoro- or2,2,2-trichloro-acetyl, benzoyl that is unsubstituted or substituted by,for example, halogen, lower alkoxy or nitro, such as benzoyl,4-chlorobenzoyl, 4-methoxybenzoyl or 4-nitrobenzoyl, loweralkoxycarbonyl, lower alkoxycarbonyl that is branched preferably at the1-position of the lower alkyl radical or that is suitably substituted atthe 1- or 2-position, for example tert-lower alkoxycarbonyl, such astert-butoxycarbonyl, arylmethoxycarbonyl having one, two or three arylradicals which are phenyl that is unsubstituted or mono- orpolyo-substituted, for example, by lower alkyl, especially tert-loweralkyl, such as tert-butyl, lower alkoxy, such as methoxy, hydroxy,halogen, such as chlorine, and/or by nitro, for examplebenzyloxycarbonyl, 4-nitrobenzyloxycarbonyl, diphenylmethoxycarbonyl,9-fluorenylmethoxycarbonyl or di(4-methoxyphenyl)methoxycarbonyl,aroylmethoxycarbonyl wherein the aroyl group is preferably benzoyl thatis unsubstituted or substituted by, for example, halogen, such asbromine, for example phenacyloxycarbonyl, 2-halo-lower alkoxycarbonyl,for example 2,2,2-trichloroethoxycarbonyl, 2-bromoethoxycarbonyl or2-iodoethoxycarbonyl, 2-(trisubstituted silyl)-lower alkoxycarbonyl, forexample 2-tri-lower alkylsilyl-lower alkoxycarbonyl, such as2-trimethylsilylethoxycarbonyl or2-(di-n-butylmethylsilyl)ethoxycarbonyl, or triarylsilyl-loweralkoxycarbonyl, for example 2-triphenylsilylethoxycarbonyl. Especiallyimportant amino- (and imino-)acyl-protecting groups are suitable organicsulfonic acid radicals, such as arylsulfonic acid radicals, especiallyphenyl- or lower alkylphenyl-sulfonyl radicals, such as benzene- ortoluene-sulfonyl, or aryl-lower alkylsulfonyl radicals, especiallyphenyl- or lower alkylphenyl-sulfonyl, such as benzyl- or4-methylbenzyl-sulfonyl; and/or suitable organic phosphoryl radicals,such as diarylphosphinyl, especially diphenylphosphinyl ([Phe]₂(P═O)--), or, most especially, di(lower alkoxy)phosphoryl, such asdiethoxyphosphoryl ([H₃ C--CH₂ O--]₂ --(P═O)--).

In an arylmethylamino group, for example a mono-, di- or, especially,tri-arylmethylamino group, the aryl radicals are especiallyunsubstituted or substituted phenyl radicals. Such groups are, forexample, benzyl-, diphenylmethyl- or, especially, trityl-amino.

In a 2-acyl-lower alk-1-enyl radical that may be used as anamino-protecting group, acyl is, for example, the corresponding radicalof a lower alkanecarboxylic acid, of a benzoic acid that isunsubstituted or substituted, for example, by lower alkyl, such asmethyl or tert-butyl, lower alkoxy, such as methoxy, halogen, such aschlorine, and/or by nitro, or especially of a carbonic acid semiester,such as a carbonic acid lower alkyl semiester. Corresponding protectinggroups are especially 1-lower alkanoyl-lower alk-1-en-2-yl, for example1-lower alkanoyl-prop-1-en-2-yl, such as 1-acetyl-prop-1-en-2-yl, orlower alkoxycarbonyl-lower alk-1-en-2-yl, for example loweralkoxycarbonyl-prop-1-en-2-yl, such as 1-ethoxycarbonyl-prop-1-en-2-yl.

A silylamino group is, for example, a tri-lower alkylsilylamino group,for example trimethylsilylamino or tert-butyl-dimethylsilylamino. It isalso possible for the silicon atom of the silylamino group to besubstituted by only two lower alkyl groups, for example methyl groups,and by the amino group or carboxy group of a second molecule of formulaI. Compounds having such protecting groups may be prepared, for example,using the corresponding chlorosilanes, such as dimethylchlorosilane, assilylating agent.

Preferred amino-protecting groups are lower alkoxycarbonyl, phenyl-loweralkoxycarbonyl, fluorenyl-lower alkoxycarbonyl, 4-loweralkylphenylsulfonyl, di(lower alkoxy)phosphoryl, 2-lower alkanoyl-loweralk-1-en-2-yl or lower alkoxycarbonyl-lower alk-1-en-2-yl, withtert-butoxycarbonyl, toluenesulfonyl and/or diethoxyphosphoryl beingespecially preferred.

Preferred are also two divalent amino-protecting groups (preferablybridging adjacent nitrogen atoms, such as N¹ and N⁵ or N¹⁰ and N¹⁴),such as unsubstituted or mono- or di-substituted methylene groups, suchas 1-lower alkoxy (for example methoxy or ethoxy)lower alkylene (forexample ethylene or 1-n-butylene), for example --C(CH₃)(OC₂ H₅)--,especially mono- or di-lower alkyl- or phenyl-methylene, for example--C(CH₃)₂ -- or --CH(-phenyl)--; methylene (--CH₂ --) is especiallypreferred.

In the starting materials of formula III, X is preferably a nucleofugalgroup, preferably arylsulfonyloxy, such as toluenesulfonyloxy, loweralkanesulfonyloxy, such as methanesulfonyloxy, or especially halogen,such as chlorine, bromine or iodine, most especially bromine or iodine.

The reaction is preferably carried out in the presence of a strong base,such as an alkali metal hydride, for example sodium hydride or potassiumhydride, or also an alkali metal amide, such as sodium amide, or analkali metal di-lower alkylamide, such as lithium diisopropylamide,especially in the presence of sodium hydride or potassium hydride, whichmay be added, for example, in the form of a dispersion in oil or afterextraction of the oil, for example with a liquid hydrocarbon, such ashexane, using the base in an equimolar amount or preferably in excessrelative to the molar amount of the compound of formula II, for examplein an amount of from 1 to 20 times the molar amount, especially from 2to 10 times the molar amount, at preferred temperatures of from -10° C.to the reflux temperature of the reaction mixture, especially fromapproximately 5 to approximately 40° C., for example at roomtemperature, or (when starting materials containing phosphoryl-protectedamino groups are used) at from 10° C. to the reflux temperature, forexample at from 20° to 80° C., in aprotic, especially polar, solvents,such as acid amides, for example dimethylformamide, diethylformamide,1,3-dimethyl-3,4,5,6-tetrahydro-2(1H)-pyrimidinone (DMPU) orhexamethylphosphoric acid triamide, aromatic hydrocarbons, such astoluene or benzene (in which case preferably in the presence of a phasetransfer catalyst, for example a tetra-lower alkylammonium halide, suchas tetra(n-butyl)ammonium bromide), or mixtures of such solvents, in thepresence or absence of a protecting gas, such as argon or nitrogen;ammonia that is formed when alkali metal amides are used as bases ispreferably removed by the application of a vacuum, for example of from0.1 to 100, especially from 0.5 to 10, torr.

It is preferred to use the compound of formula III also in an equimolaramount or in excess relative to the compound of formula II, especiallyin an amount that is from 1 to 20 times the molar amount, especiallyfrom 2 to 10 times the molar amount, relative to the compound of formulaII.

When the protecting group at the nitrogen atom to be alkylated by R₁ isan organic sulfonic acid radical, such as arylsulfonyl, especiallyphenylsulfonyl or lower alkylphenylsulfonyl, such as benzene- ortoluene-sulfonyl, or aryl-lower alkylsulfonyl, especially phenyl- orlower alkylphenylsulfonyl, such as benzyl- or 4-methylbenzyl-sulfonyl,the alkylation with a compound of formula III can preferably be carriedout in the presence of relatively weak bases, such as especially metalhydroxides or carbonates, such as especially alkali metal hydroxides,for example sodium or potassium hydroxide, or in the presence ofalkaline earth metal carbonates or alkali metal carbonates, for examplesodium or potassium carbonate, preferably in the last-mentionedsolvents, especially in halogenated hydrocarbons, such asdichloromethane or chloroform, and most especially in carboxylic acidamides, such as dimethylformamide or dimethylacetamide, and at thetemperatures indicated and preferably under a protecting gas, such asnitrogen or argon.

The following applies in respect of the subsequent removal of theprotecting groups:

A protected amino or imino group is freed in a manner known per se and,depending on the nature of the protecting groups, by different methods,preferably by means of solvolysis or selective reduction, for example asdescribed in the standard works mentioned at the beginning. Loweralkoxycarbonylamino, such as tert-butoxycarbonylamino,arylmethoxycarbonylamino, such as (unsubstituted or substituted)benzyloxycarbonylamino, or di(lower alkoxy)phosphoryl can be removed inthe presence of acids, for example mineral acids, e.g. a hydrogenhalide, such as hydrogen chloride or hydrogen bromide, or of sulfuric orphosphoric acid, preferably in the presence of hydrogen chloride, inpolar solvents, such as water, alcohols, such as lower alkanols, e.g.methanol or ethanol, a carboxylic acid, such as acetic acid, or ethers,preferably cyclic ethers, such as tetrahydrofuran or dioxane (preferredin the case of di-lower alkoxyphosphoryl removal), or mixtures of two ormore of the mentioned solvents, especially in aqueousalcoholicsolutions, such as water/methanol mixtures; and 2-halo-loweralkoxycarbonylamino (optionally after conversion of a 2-bromo-loweralkoxycarbonylamino group into a 2-iodo-lower alkoxycarbonylaminogroup), aroylmethoxycarbonylamino or 4-nitrobenzyloxycarbonylamino canbe removed, for example, by treatment with a suitable reducing agent,such as zinc in the presence of a suitable carboxylic acid, such asaqueous acetic acid. Aroylmethoxycarbonylamino can be removed also bytreatment with a nucleophilic, preferably salt-forming reagent, such assodium thiophenolate, and 4-nitrobenzyloxycarbonylamino can be removedalso by treatment with an alkali metal dithionite, for example sodiumdithionite. Unsubstituted or substituted diphenylmethoxycarbonylamino,tert-lower alkoxycarbonylamino or 2-(trisubstituted silyl)-loweralkoxycarbonylamino, such as 2-tri-lower alkylsilyl-loweralkoxycarbonylamino, can be removed by treatment with a suitable acid,for example formic acid or trifluoroacetic acid, and unsubstituted orsubstituted triarylmethylamino or formylamino can be removed, forexample, by treatment with an acid, such as a mineral acid, for examplehydrochloric acid, or an organic acid, for example formic acid, aceticacid or trifluoroacetic acid, in the absence or presence of water, andan amino group protected in the form of silylamino can be freed, forexample, by means of hydrolysis or alcoholysis. An amino group protectedby 2-haloacetyl, for example 2-chloroacetyl, can be freed by treatmentwith thiourea in the presence of a base, or with a thiolate salt, suchas an alkali metal thiolate, of thiourea, and subsequent solvolysis,such as alcoholysis or hydrolysis, of the resulting substitutionproduct. An amino group protected by 2-(trisubstituted silyl)-loweralkoxycarbonyl, such as 2-tri-lower alkylsilyl-lower alkoxycarbonyl, canbe converted into free amino (or imino) also by treatment with a salt ofhydrofluoric acid that yields fluoride anions, such as an alkali metalfluoride, for example sodium or potassium fluoride, in the absence orpresence of a macrocyclic polyether ("crown ether"), or with a fluorideof an organic quaternary base, such as tetra-lower alkylammoniumfluoride or tri-lower alkylaryl-lower alkylammonium fluoride, forexample tetraethylammonium fluoride or tetrabutylammonium fluoride, inthe presence of an aprotic, polar solvent, such as dimethyl sulfoxide orN,N-dimethylacetaminde. Likewise, silyl, such as trimethylsilyl, bondeddirectly to a hetero atom, such as nitrogen, can be removed by means offluoride ions. An amino group protected by diarylphosphinyl, such asdiphenylphosphinyl, can be freed in the presence of a Lewis acid,especially boron trifluoride etherate, such as boron trifluoride ethyletherate or methyl etherate, in suitable solvents or solvent mixtures,for example alcohols, such as methanol or ethanol, halogenatedhydrocarbons, such as chloroform or methylene chloride, ethers, such asdimethyl or diethyl ether, or especially mixtures thereof, such asmethanol/methylene chloride/dimethyl or diethyl ether, at preferredtemperatures of from -10° C. to the respective reflux temperature,especially from 0° C. to room temperature, preferably under a protectinggas, such as N₂.

An amino group protected in the form of a sulfonamide is preferablyfreed by acid hydrolysis, for example in the presence of a mineral acid,such as especially a hydrohalic acid, such as hydrobromic acid, in analcohol, especially an aryl alcohol, such as phenol, in the presence orabsence of a carboxylic acid, such as a lower alkanoic acid, for exampleacetic acid, at preferred temperatures of from 60° C. to the refluxtemperature, or by acid hydrolysis with concentrated sulfuric acid.

An amino group protected by di(lower alkoxy)phosphoryl is preferablyfreed by acid hydrolysis, for example in the presence of a hydrogenhalide, such as hydrogen bromide or especially hydrogen chloride (whichis preferably introduced in gaseous form), in an ether, especially acyclic ether, such as tetrahydrofuran, at preferred temperatures of from-10° C. to the reflux temperature of the reaction mixture in question,for example at from approximately 0° C. to approximately roomtemperature.

Nitrogen atoms protected by a divalent protecting group bridging twoadjacent nitrogen atoms are preferably freed by acidolytic cleavage, forexample with mineral acids, such as hydrohalic acids, for examplehydrochloric or hydrobromic acid, or under milder conditions similar toKnoevenagel conditions, for example with malonic acid or cyanoaceticacid in the presence of a tertiary nitrogen base, such as pyridine (see,for example, Nagarajan et at., J. Org. Chem. 50, 5735-5737 (1985)).

The temperatures at which the protected functional groups are freed arepreferably from -80° C. to reflux temperature, especially preferablyfrom -20° to 50° C. or from 80° to 110° C., for example from 0° to 35°C., such as in the range of from 0° C. to room temperature, orapproximately at reflux temperature.

When several protected functional groups are present, the protectinggroups may, if desired, be so selected that it is possible to removemore than one such group simultaneously. Conversely, the groups may alsobe so selected that they are not all removed simultaneously but can beremoved in a desired sequence, in which case the correspondingintermediates are obtained.

The introduction of amino- and/or imino-protecting groups that isnecessary for the preparation of protected starting materials of formulaII is effected in a manner known per se, for example as described in theabove-mentioned standard works, and may be carried out stepwise or,preferably, in a single procedure.

For the introduction of the acyl protecting group of a carbonic acidsemiester, such as lower alkoxycarbonyl, there are suitable especiallysymmetrical or mixed carbonic acid anhydrides, such as di-lower alkyldicarbonate, for example di-tert-butyl dicarbonate, or loweralkoxycarboxylic acid azides, such as tert-butoxycarboxylic acid azide,or other activated carbonic acid semiester derivatives, such asimidazolides, for example lower alkoxy-, such as tert-butoxy-carboxylicacid 1-imidazolide, or especially2-(tert-butoxycarbonyl-oxyimino)-2-phenylacetonitrile.

For the introduction of organic sulfonic acid radicals, such asarylsulfonic acid radicals, especially phenyl- or loweralkylphenyl-sulfonyl radicals, such as benzene- or toluenesulfonyl, oraryl-lower alkylsulfonyl radicals, especially phenyl- or loweralkylphenylsulfonyl, such as benzyl- or 4-methylbenzyl-sulfonyl, thereare suitable especially corresponding sulfonyl halides, such as sulfonylchlorides or bromides, for example toluenesulfonic acid chloride.

For the introduction of suitable organic phosphoryl radicals, such asdiarylphosphinyl, especially diphenylphosphinyl, or more especiallydi(lower alkoxy)phosphoryl, such as diethoxyphosphoryl, there issuitable especially reaction with corresponding phosphoryl halides, forexample chlorides, such as diphenylphosphinyl chloride (see Osborn, H.M. I., et al., Synlett 2, 145-147 (1994)), or iodides, such as di-loweralkoxyphosphoryl iodide (which can be prepared, for example,electrochemically in situ in acetonitrile on platinum electrodes insupporting electrolytes, such as tetra-lower alkylammonium halides, forexample tetraethylammonium bromide, see J. Gen. Chem. (USSR) 62, 370(1992)); or (for the introduction especially of di(loweralkoxy)phosphoryl) from the corresponding phosphites, such as all(loweralkyl) phosphite, especially diethyl phosphite, under substantiallyanhydrous conditions, for example by phase transfer catalysis in thepresence of a phase transfer catalyst, for example of a tetra-loweralkylammonium halide, such as tetra(n-butyl)ammonium bromide; in thepresence (in each case preferably in excess, for example in a 2- to20-fold molar excess, relative to the base to be protected) of adehydrating inorganic salt, such as potassium carbonate, and of a basethat can be convened by the phase transfer catalyst from the solid forminto the organic solution, such as potassium hydrogen carbonate; insuitable organic solvents or solvent mixtures, such as halogenatedhydrocarbons, for example methylene chloride or carbon tetrachloride, ormixtures thereof; at preferred temperatures of from 0° to 40° C., forexample at from approximately 10° to approximately 30° C. (see J. Org.Chem. 56, 4904-4907 (1991)); reaction without a phase transfer catalystalso being possible.

For the introduction of divalent amino-protecting groups, such asunsubstituted or monoor di-substituted methylene groups, such as 1-loweralkoxy (for example methoxy or ethoxy)-lower alkylene (for exampleethylene or 1-n-butylene), for example --C(CH₃)(OC₂ H₅)--, conventionalmethods are used. Especially for the introduction of mono- or di-loweralkyl- or phenyl-methylene, for example --C(CH₃)₂ -- or --CH(-phenyl)--,especially --CH₂ --, there are suitable corresponding aldehydes orketones in which there is an oxo group in place of the two bondsindicated in the formulae shown above, for example benzaldehyde orespecially acetone or most especially formaldehyde.

The introduction is carded out under customary conditions, preferably insolvents, such as carboxylic acid amides, for example dimethyl- ordiethyl-formamide, in chlorinated hydrocarbons, such as carbontetrachloride, chloroform or methylene chloride, or in ethers, such ascyclic ethers, for example tetrahydrofuran, or mixtures thereof, in thecase of the introduction of sulfonyl radicals additionally in thepresence of water (biphasic system) and in the case of the introductionof unsubstituted, mono- or di-substituted methylene groups also ifdesired additionally or exclusively in the presence of water, it beingpossible in each case for the water to contain a base, such as an alkalimetal hydroxide, such as sodium or potassium hydroxide, if necessaryunder a protecting gas, such as nitrogen or argon, and if necessary inthe presence of bases, such as tertiary nitrogen bases, for exampletriethylamine, pyridine or 4-dimethylaminopyridine, or of morpholine, orhydroxides, such as ammonium hydroxide or alkali metal hydroxides, forexample sodium or potassium hydroxide.

Preferred temperatures are from -10° to 50° C., especially from 0° to30° C.

Where indicated, the conditions mentioned specifically in eachparticular case are preferred.

Starting materials of formula II can preferably be prepared from acompound of formula IV by, especially, reacting a compound of formula IVthat is protected analogously to a compound of formula II (as describedabove) and wherein all four nitrogen atoms are protected but a hydrogenis bonded to the nitrogen to be reacted at least at the position atwhich R₂ is to be introduced, with a compound of formula IX

    R.sub.2 --X                                                (IX),

wherein R₂ is as defined for compounds of formula I and X is as definedunder formula III, under conditions analogous to those described forprocess a), and obtaining the corresponding compound of formula II by,if necessary, removing some of the protecting groups at the terminalnitrogen to which R₁ is to be bonded, or by removing all the protectinggroups and then re-introducing protecting groups, as described forcompounds of formula II. A compound of formula II protected as describedabove may also be isolated as a secondary product of process b).

Process variant a) is suitable especially for the introduction ofradicals R₁ and R₂ that are different from each other, in which casecorresponding non-symmetrical compounds of formula I are obtained, orfor the introduction of sterically hindered radicals, such as isobutylor isopropyl.

Process b) (alkylation): In the starting materials of formula IV, allthe nitrogen atoms are preferably in mono-protected form, so that thehydrogen that is to be replaced by R₁ and R₂ is still present; in thatcase the 4 nitrogen atoms are each bonded to a protecting group insteadof to one of the 4 hydrogen atoms shown in formula IV that are not to bereacted.

In compounds of formula V, X is as defined above for compounds offormula III.

The protecting groups, the process conditions for the reaction,including the molar ratios of the starting materials, the removal ofprotecting groups and the introduction of protecting groups arepreferably analogous to those described under process a), wherecompounds of formula IV are used instead of compounds of formula II andcompounds of formula V are used instead of compounds of formula III.

The reaction is suitable especially for the introduction of two radicalsR_(l) and R₂ that are the same. In that case, symmetrically substitutedcompounds of formula I are formed.

Compounds of formula II can be obtained as secondary product in thisreaction, in some cases in a relatively large amount, and can beisolated and used in process a). This is a preferred method of synthesisfor the preparation of non-symmetrical compounds of formula I.

The unprotected starting material of formula IV is known, is obtainablee.g. by reaction of trans-1,4-dichloro-2-butene with 3-aminopropylamineprotected at an amino nitrogen and subsequent removal of the protectinggroups or is available commercially (e.g. from Carbolabs, Inc., NewHaven, Conn., USA; or Ames Laboratories, Inc., Milford, Conn., USA).

Process c) (reduction of amides): In the diamides of formula VI whereinR₁ ' and R₂ ' are each independently of the other the radicals, minusthe above-mentioned methylene group, which are complementary to thelast-defined radicals R₁ and R₂, and any functional groups that are notto take part in the reaction are, if necessary, in protected form, R₁'--(C═O)-- and R₂ '--(C═O)-- are each independently of the otherespecially unsubstituted C₁ -C₆ alkyl-(C═O)-- (yields a C₁ -C₇ alkylradical R₁ or R₂ bonded via methylene), which may, however, also besubstituted by fluorine, especially by up to three fluorine atoms; C₂-C₆ alkenyl-(C═O)-- (yields a C₃ -C₇ alkenyl radical R₁ or R₂ bonded viamethylene), preferably C₂ -C₃ alkenyl-(C═O)--, e.g. vinyl-(C═O)-- orallyl-(C═O)--; C₂ -C₆ alkynyl-(C═O)-- (yields a C₃ -C₇ alkynyl radicalR₁ or R₂ bonded via methylene), especially C₂ -C₃ -alkynyl-(C═O)--, e.g.ethynyl or propyn-1-yl; or cycloalkyl-C₁ -C₆ alkyl-(C═O)-- (yields acycloalkyl-C₂ -C₇ alkyl radical R₁ or R₂ bonded via a methylene group)or cycloalkyl-(C═O)--, wherein cycloalkyl is especially a radical havingfrom 3 to 5 carbon atoms, especially cyclopropyl or also cyclobutyl, andthe C₁ -C₆ alkyl radical (if present) is preferably a radical defined asabove, especially C₁ -C₂ alkyl, for example methyl or ethyl, especiallymethyl, e.g. cyclopropyl-methyl-(C═O)--, cyclobutyl-(C═O)-- or mostespecially cyclopropyl-(C═O)--.

The two radicals R₁ ' and R₂ ' may be different or, especially, thesame. The resulting radicals R₁ and R₂ are without primary branching atthe bonding carbon atom.

The selective reduction of the two amide groups may be carded outsimultaneously or stepwise. "Selective" means that the reducing agentdoes not at the same time reduce the central double bond and any furtherolefinic bonds or carbon-carbon triple bonds that are present, butprotecting groups removable under the corresponding conditions can beremoved virtually at the same time (i.e. in the same procedure) as theamide group reduction. Suitable selective reducing agents are especiallycertain complex hydrides, such as NaAl(OCH₂ CH₂ OCH₃)₂ H₂ (=sodiumdihydro-bis(2-methoxyethoxy)aluminate ), which can be added in the formof a solution in an aromatic hydrocarbon, such as toluene, or lithiumaluminium hydride, which are preferably used in ethers as solvents, suchas cyclic ethers, for example dioxane or tetrahydrofuran, or in di-loweralkyl ethers, such as diethyl ether, with heating, for example at refluxtemperature (especially in the case of diethyl ether or tetrahydrofuran)or at temperatures below the reflux temperature, for example at from 20°to 35° C.

Since secondary reactions may occur in this process, processes a), b) ord) are preferred.

The protecting groups and their introduction and removal are preferablyas described under process a). Preferred protecting groups are thementioned divalent protecting groups selected from unsubstituted, mono-or di-substituted methylene, especially methylene, as a bridge betweenN¹ and N⁵ as well as between N¹⁰ and N¹⁴, which are introduced asdescribed above.

The starting materials of formula VI can be prepared in particular,compounds of formula VI' ##STR8## wherein R₁ ' and R₂ ' are as definedand Y is one of the above-mentioned divalent amino-protecting groups,especially methylene, (these are preferred compounds of formula VI), canbe prepared from corresponding precursors wherein there is a hydrogenatom in place of one or both of the radicals R₁ --(C═O)-- and R₂--(C═O)-- and the remaining radicals are as last defined, by reactingthose precursors with a carboxylic acid of formula X

    R.sub.1 '--(C═O)--OH                                   (X)

and (especially when R₁ ' and R₂ ' in compounds of formula VI aredifferent) then with a carboxylic acid of formula XI

    R.sub.2 '--(C═O)--OH                                   (XI),

wherein R₁ ' and R₂ ' are each as defined for compounds of formula VI,or preferably with reactive derivatives of the compounds of formula Xand, where appropriate, of formula XI, under customary conditions forthe preparation of acid amides.

Reactive acid derivatives are especially the corresponding symmetricalacid anhydrides of the formulae R₁ '--(C═O)--O--(C═O)--R₁ ' and R₂'--(C═O)--O--(C═O)--R₂ ', or also asymmetrical acid anhydrides,especially the corresponding acid chlorides, acid azides or mixedanhydrides with acetic acid, or also the hydroxysuccinimide derivatives,all of which are known, can be prepared according to known processes orare available commercially.

Derivatives of carboxylic acids of formula X or XI, which are used asacylating agents, can also be formed in situ. For example,N,N'-disubstituted amidino esters can be formed in situ by reacting themixture of the starting material of formula IV and the acid used asacylating agent in the presence of a suitable N,N'-disubstitutedcarbodiimide, for example N,N'-cyclohexylcarbodiimide. Furthermore,amino or amido esters of the acids used as acylating agent can be formedin the presence of the starting material of formula IV that is to beacylated, by reacting the mixture of the corresponding acid and aminostarting materials in the presence of an N,N'-disubstitutedcarbodiimide, for example N,N'-dicyclohexylcarbodiimide, and of anN-hydroxy-imide, for example N-hydroxysuccinimide, in the absence orpresence of a suitable base, for example 4-dimethylamino-pyridine.Moreover, activation can be carried out in situ by reaction withN,N,N',N'-tetraalkyluronium compounds, such asO-benzotriazol-1-yl-N,N,N',N'-tetramethyluronium hexafluorophosphate,O-(1,2-dihydro-2-oxo- 1-pyridyl)-N,N,N',N'-tetramethyluroniumtetrafluoroborate or O-(3,4-dihydro-4-oxo-1,2,3-benzo-triazolin-3-yl)-N,N,N',N'-tetramethyluroniumtetrafluoroborate. Finally, phosphoric acid anhydrides of the carboxylicacids of formulae X or XI can be prepared in situ by reacting analkylphosphoric acid amide, such as hexamethylphosphoric acid triamide,in the presence of a sulfonic acid anhydride, such as 4-toluenesulfonicacid anhydride, with a salt, such as a tetrafluoroborate, for examplesodium tetrafluoroborate, or with another derivative ofhexamethylphosphoric acid triamide, such asbenzotriazol-1-yl-oxy-tris(dimethylamino)phosphonium hexafluoride,preferably in the presence of an additive, such asN-hydroxybenzotriazole.

The acylation is preferably carried out in an inert, aprotic, preferablyanhydrous solvent or solvent mixture, for example in a carboxylic acidamide, for example formamide or dimethylformamide, a halogenatedhydrocarbon, for example methylene chloride, carbon tetrachloride orchlorobenzene, a ketone, for example acetone, a cyclic ether, forexample tetrahydrofuran, an ester, for example ethyl acetate, or anitrile, for example acetonitrile, or in a mixture thereof, whereappropriate at reduced or elevated temperature, for example in atemperature range of from approximately -40° C. to approximately +100°C., preferably from approximately -10° C. to approximately +50° C., inthe case where arylsulfonyl esters are used at approximately from +100°C. to +200° C., and where appropriate under an inert gas atmosphere, forexample under a nitrogen or argon atmosphere.

Aqueous, for example alcoholic, solvents, e.g. ethanol, or aromaticsolvents, e.g. benzene or toluene, are also possible.

The reaction is preferably carried out in the presence of tertiarynitrogen bases, such as pyridine or triethylamine, also4-dimethylaminopyridine or N-methyl-morpholine. The mentioned bases,especially pyridine, may be present instead of a solvent; that ispreferably the case when symmetrical anhydrides are used as reactiveacid derivatives (those anhydrides are then used preferably in a molarexcess relative to the amine to be acylated, preferably in a 1.2- to10-fold excess).

The precursors of the compounds of formula VI', wherein a hydrogen atomis present in place of one or both of the radicals R₁ --(C═O)-- and R₂--(C═O)-- and the remaining radicals are as defined, are prepared, forexample, by introducing the divalent amino-protecting groups Y into acompound of formula IV under conditions such as those described in themore detailed description of process variant a), preferably usingformaldehyde in order to introduce two methylene protecting groups, asdescribed therein.

Process d) (nucleophilic substitution): In compounds of formula VII, adivalent protecting group Y is especially an unsubstituted or mono- ordi-substituted methylene group, such as 1-lower alkoxy (for examplemethoxy or ethoxy)-lower alkylene (for example ethylene or1-n-butylene), e.g. --C(CH₃)(OC₂ H₅)--, especially e.g. --C(CH₃)₂ -- or--CH(-phenyl)--; methylene (--CH₂ --) is especially preferred.

In compounds of formula VIII, Q and Q' are nucleofugal leaving groups,preferably as defined above for X in compounds of formula III,especially halogen, such as bromine or iodine. Q and Q' are preferablyidentical.

The compound of formula VII is preferably used in a two-fold or greater,especially a 2- to 10-fold, molar excess relative to the compound offormula VIII, which yields primarily Y-protected compounds of formula Iwherein R₁ and R₂ are identical. It is, however, also possible tosubstitute first the radical Q and then, in the monohexahydropyrimidineintermediate that is obtainable, which can be isolated or processedfurther directly in situ, to react the radical Q' with a differentcompound of formula VII (wherein Rx and/or Y have a different meaningthan in the compound of formula VII that has already been reacted). Forexample, it is also possible to obtain compounds of formula I wherein R₁and R₂ are different from each other. For that purpose an excess is notabsolutely necessary, that is to say each of the two compounds offormula VII can be used, for example, in a 1- to 10-fold excess,especially in a 1- to 5-fold excess, relative to the compound of formulaVIII.

The reaction is carried out under the conditions customary for anucleophilic substitution, preferably in aprotic solvents, such asketones, for example a di-lower alkyl ketone, such as acetone, nitriles,for example a lower alkylnitrile, such as acetonitrile, carboxylic acidamides, for example a di-lower alkyl-lower alkanoylamide, such asdimethylformamide or dimethylacetamide, di-lower alkyl sulfoxides, suchas dimethyl sulfoxide, in DMPU, hexamethylphosphoric acid triamide orethers, such as di-lower alkyl ethers, for example diethyl ether, orcyclic ethers, such as tetrahydrofuran or dioxane, or also in proticsolvents, such as alcohols, especially lower alkanols, for examplemethanol or ethanol, or mixtures of two or more of the mentionedsolvents.

The starting materials of formula VII are preferably prepared byring-closure of the corresponding N-Rx-substituted trimethylenediamineswith the corresponding aldehydes or ketones suitable for theintroduction of Y (such as, especially, formaldehyde or acetone), whichare used for the introduction of the divalent protecting groups Y,preferably under conditions described as preferred in connection withthe introduction of such protecting groups under process a), for exampleanalogously to the process described in Okada, J., et al., Chem. Pharm.Bull. 28(11), 3310-3314 (1980).

Conversion of salts and separation of isomers

The conversion of a salt of a compound of formula I into a differentsalt is carded out, for example, in solvents, especially in organicsolvents, more especially in polar organic solvents, very especially inesters, for example lower alkanoyl-lower alkyl esters, such as ethylacetate, in amides, for example N,N-di-lower alkyl-lower alkanoylamides,such as dimethylformamide, in alcohols, for example hydroxy-loweralkanes, such as methanol, ethanol, ethylene glycol or glycerol, or arylalcohols, such as phenols, for example phenol, or in dimethyl sulfoxide,in the absence or presence of water, preferably in the presence ofwater, or in water itself. Special preference is given to reaction inalcohols, such as the last-mentioned hydroxy-lower alkanes, in mixturesof such alcohols and water, or in water itself.

The reaction is carded out, for example, in free solution, but it mayalso be effected over chromatographic columns, for example by gelfiltration, over ion exchangers or by means of semi-permeable membranesby osmotic processes, for example by dialysis.

The reaction is carried out at temperatures from the freezing point tothe boiling point of the solutions in question, preferably at from 0° to50° C., especially at from 20° to 40° C., for example at roomtemperature, in the presence or absence of a protecting gas, such asnitrogen or argon.

The compounds of formula I and the salt-forming acid are used insuitable molar ratios, or the acid is employed in excess. Preferably,the individual components are used in the molar ratio that correspondsto the ratio of the molarity of the base of formula I and the acid inthe resulting salts.

The salts that are formed precipitate, for example, by themselves, insome cases only after cooling, or they are precipitated by the additionof solvents, especially of non-polar solvents, for example ethers, suchas diethyl ether, or of water, and/or are obtained by partial orcomplete concentration by evaporation.

The reaction may also be effected via the free bases of formula I, whichare prepared, for example, by converting the acid salt of a base offormula I, with a first acid, used as starting material into the freebase with the aid of a base, for example a hydroxy base, such as analkali metal hydroxide, for example NaOH or KOH, or with an OH⁻ -chargedion exchanger, such as ®Amberlite-IRA-400 in the OH⁻ form, in aqueoussolution in the presence or absence of an organic solvent, as definedabove; the subsequent conversion of the free base is carried out, forexample, as described above.

The free bases of the compounds of formula I are preferably prepared asjust described, also by chromatography, for example by gel filtration,or over ion exchangers.

Mixtures of isomers obtainable according to the invention can beseparated in a manner known per se into the individual isomers;diastereoisomers can be separated, for example, by partitioning betweenpolyphasic solvent mixtures, recrystallisation and/or chromatographicseparation, for example over silica gel, and racemates can be separated,for example, by the formation of salts with optically pure salt-formingreagents and separation of the mixture of diastereoisomers soobtainable, for example by means of fractional crystallisation, or bychromatography over optically active column materials.

General process conditions

The following applies in general to all processes mentioned hereinbeforeand hereinafter:

Unless a specific method of synthesis is indicated for startingmaterials, the starting materials are known, can be prepared accordingto processes known per se and/or are available commercially.

In view of the close relationship between the compounds of formula I andtheir salts and starting materials (starting materials andintermediates) in free form and in the form of their salts, anyreference hereinbefore and hereinafter to the free compounds or theirsalts is to be understood as meaning also the corresponding salts orfree compounds, respectively, where appropriate and expedient.

All the above-mentioned process steps can be carried out under reactionconditions that are known per se, preferably those mentionedspecifically, in the absence or, customarily, in the presence ofsolvents or diluents, preferably solvents or diluents that are inerttowards the reagents used and are solvents therefor, in the absence orpresence of catalysts, condensation agents or neutralising agents, forexample ion exchangers, such as cation exchangers, e.g. in the H⁺ form,depending on the nature of the reaction and/or of the reactants atreduced, normal or elevated temperature, for example in a temperaturerange of from approximately -100° C. to approximately 190° C.,preferably from approximately -80° C. to approximately 150° C., forexample at from -80° to -60° C., at room temperature, at from -20° to40° C. or at reflux temperature, under atmospheric pressure or in aclosed vessel, where appropriate under pressure, and/or in an inertatmosphere, for example under an argon or nitrogen atmosphere.

At all stages of the reactions, mixtures of isomers that are formed canbe separated into the individual isomers, for example diastereoisomersor enantiomers, or into any desired mixtures of isomers, for exampleracemates or mixtures of diastereoisomers, for example analogously tothe methods described under "Conversion of salts and separation ofisomers".

The solvents from which those solvents that are suitable for anyparticular reaction may be selected include, for example, water, esters,such as lower alkyl-lower alkanoates, for example ethyl acetate, ethers,such as aliphatic ethers, for example diethyl ether, or cyclic ethers,for example tetrahydrofuran, liquid aromatic hydrocarbons, such asbenzene or toluene, alcohols, such as methanol, ethanol or 1- or2-propanol, nitriles, such as acetonitrile, halogenated hydrocarbons,such as methylene chloride, acid amides, such as dimethylformamide,bases, such as heterocyclic nitrogen bases, for example pyridine,carboxylic acid anhydrides, such as lower alkanoic acid anhydrides, forexample acetic anhydride, cyclic, linear or branched hydrocarbons, suchas cyclohexane, hexane or isopentane, or mixtures of those solvents, forexample aqueous solutions, unless otherwise indicated in the descriptionof the processes. Such solvent mixtures may also be used in working up,for example by chromatography or partitioning.

The compounds, including their salts, may also be obtained in the formof hydrates, or their crystals may, for example, include the solventused for crystallisation.

If necessary, protected starting materials may be used in all processsteps and the protecting groups may be removed at suitable stages of thereaction.

The invention relates also to those forms of the process in which acompound obtainable as intermediate at any stage of the process is usedas starting material and the remaining process steps are carried out, orin which a starting material is formed under the reaction conditions oris used in the form of a derivative, for example in protected form or inthe form of a salt, or a compound obtainable by the process according tothe invention is produced under the process conditions and processedfurther in situ. In the process of the present invention there arepreferably used those starting materials which result in the compoundsof formula I described at the beginning as being especially valuable.Special preference is given to reaction conditions that are analogous tothose mentioned in the Examples.

Pharmaceutical compositions and processes

The present invention relates also to pharmaceutical compositions thatcomprise compounds of formula I as active ingredient. Compositions forenteral, especially oral, and parenteral administration are especiallypreferred. The compositions comprise the active ingredient on its ownor, preferably, together with a pharmaceutically acceptable carrier. Thedose of active ingredient depends on the disease to be treated, and onthe species, its age, weight and individual condition, and on the modeof administration.

Preference is given to a pharmaceutical composition that is suitable foradministration to a warm-blooded animal, especially a human, sufferingfrom a disorder that is responsive to a reduction in the intracellularconcentrations of natural polyamines, such as especially putrescine,spermidine and spermine, especially one of the above-mentioneddisorders, for example tumour disorders or protozoal infections, whichcomposition comprises a compound of formula I or a salt thereof in anamount that is effective in the treatment of the mentioned disorders,together with at least one pharmaceutically acceptable carrier.

The pharmaceutical compositions comprise from approximately 5% toapproximately 95% active ingredient, dosage forms that are in singledose form preferably comprising from approximately 20% to approximately90% active ingredient, and dosage forms that are not in single dose formpreferably comprising from approximately 5% to approximately 20% activeingredient. Unit dose forms, such as dragees, tablets or capsules,comprise from approximately 0.05 g to approximately 1.5 g of activeingredient.

The pharmaceutical compositions of the present invention are preparedfor example by means of conventional mixing, granulating, confectioning,dissolving or lyophilising processes. For example, pharmaceuticalcompositions for oral administration can be obtained by combining theactive ingredient with one or more solid carriers, optionallygranulating a resulting mixture, and processing the mixture or granules,if desired and/or appropriate, by the addition of additional excipients,to form tablets or dragee cores.

Suitable carders are especially fillers, such as sugars, for examplelactose, saccharose, mannitol or sorbitol, cellulose preparations and/orcalcium phosphates, for example tricalcium phosphate or calcium hydrogenphosphate, and also binders, such as starches, for example corn, wheat,rice or potato starch, methylcellulose, hydroxypropylmethylcellulose,sodium carboxymethylcellulose and/or polyvinylpyrrolidone, and/or, ifdesired, disintegrators, such as the above-mentioned starches and alsocarboxymethyl starch, cross-linked polyvinylpyrrolidone, or alginic acidor a salt thereof, such as sodium alginate. Additional excipients areespecially flow conditioners and lubricants, for example silicic acid,talc, stearic acid or salts thereof, such as magnesium or calciumstearate, and/or polyethylene glycol, or derivatives thereof.

Dragee cores can be provided with suitable, where appropriate entericcoatings, there being used inter alia concentrated sugar solutions,which may comprise gum arabic, talc, polyvinylpyrrolidone, polyethyleneglycol and/or titanium dioxide, or coating solutions in suitable organicsolvents or solvent mixtures or, for the preparation of entericcoatings, solutions of suitable cellulose preparations, such asacetylcellulose phthalate or hydroxypropylmethylcellulose phthalate.Colourings or pigments may be added to the tablets or dragee coatings,for example for identification purposes or to indicate different dosesof active ingredient.

Pharmaceutical compositions for oral administration are also hardgelatin capsules, and soft sealed capsules consisting of gelatin and aplasticiser, such as glycerol or sorbitol. The hard gelatin capsules maycontain the active ingredient in the form of granules, for example inadmixture with fillers, such as corn starch, binders and/or glidants,such as talc or magnesium stearate, and, where appropriate, stabilisers.In soft capsules the active ingredient is preferably dissolved orsuspended in suitable liquid excipients, for example aqueous solutions,fatty oils, such as sesame oil, fatty acid esters of the ethylene glycolor propylene glycol type, such as ®Lauroglycol (1,2-propylene glycolmonolaurate, as a mixture of the two constitutional isomers; GattefosseS. A., Saint Priest, France), ®Gelucire (glycerides and partialpolyglycerides of fatty acids; Gattefosse S. A., Saint Priest, France)or sesame oil, paraffin oil or liquid polyethylene glycols, such as PEG300 or 400 (Fluka, Switzerland), it likewise being possible to addstabilisers or pharmaceutically acceptable detergents.

Other oral dosage forms are, for example, syrups prepared in customarymanner which comprise the active ingredient, for example, in dispersedform and in a concentration of approximately from 5% to 20%, preferablyapproximately 10% or in a similar concentration that provides a suitablesingle dose when administered, for example, in a measure of 5 or 10 ml.Also suitable are, for example, powdered or liquid concentrates for thepreparation of shakes, for example in milk. Such concentrates may alsobe packed in single dose quantities.

Suitable rectally administrable pharmaceutical compositions are, forexample, suppositories that consist of a combination of the activeingredient with a suppository base. Suitable suppository bases are, forexample, natural or synthetic triglycerides, paraffin hydrocarbons,polyethylene glycols or higher alkanols.

For parenteral administration there are suitable, especially, aqueoussolutions of an active ingredient in water-soluble form, for example inthe form of a water-soluble salt, or aqueous injection suspensions thatcomprise viscosity-increasing substances, for example sodiumcarboxymethylcellulose, sorbitol and/or dextran, and, if desired,stabilisers. The active ingredient, where appropriate together withexcipients, can also be in the form of a lyophilisate and be made into asolution prior to parenteral administration by the addition of suitablesolvents.

Solutions used, for example, for parenteral administration can also beused as infusion solutions.

The invention relates also to a process or a method for the treatment ofthe abovementioned pathological conditions, especially those which areresponsive to a reduction in the intracellular concentration ofpolyamines. The compounds of the present invention can be administeredprophylactically or therapeutically, preferably in an amount that iseffective against the mentioned disorders, to a warm-blooded animal, forexample a human, requiring such treatment, the compounds preferablybeing used in the form of pharmaceutical compositions. For a body weightof approximately 70 kg, a daily dose of from 1 mg to 8000 mg, forexample from approximately 0.1 g to approximately 7 g, preferably fromapproximately 0.5 g to approximately 5 g, of a compound of the presentinvention is administered.

The Examples which follow serve to illustrate the invention, but they donot limit the scope thereof in any way.

Temperatures are given in degrees Celsius (°C). Where no temperature isgiven, the reaction is carried out at room temperature. The R_(f)values, which indicate the relationship between the seepage propagationof the substance in question and the seepage propagation of the eluantfront, are determined on thin-layer silica gel plates by thin-layerchromatography (TLC).

The ratio of solvents and eluants to one another is always given inparts by volume, unless indicated otherwise.

The other short names and abbreviations used have the followingmeanings:

    ______________________________________                                        BOC              tert-butoxycarbonyl                                          DMF              dimethylformamide                                            h                hour(s)                                                      min.             minute(s)                                                    THF              tetrahydrofuran                                              ______________________________________                                    

The values for proton nuclear resonance spectroscopy (¹ H-NMR) are givenin ppm (parts per million) based on tetramethylsilane as internalstandard. s=singlet, d=doublet, t=triplet, q=quartet, m=multiplet,dd=double doublet.

EXAMPLE 1 (E)-1,14Di-propyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

A mixture of 1.26 g (1.84 mmol) of(E)-1,14-di-propyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 19 ml of 3N methanolic hydrochloric acid is stirred for 15 h at roomtemperature. The reaction mixture is then diluted with 19 ml of diethylether, the mixture is filtered, and the filtration residue is washedwith diethyl ether. Drying under a high vacuum at 100° C. yields thecrystalline title compound (m.p. >260° C.). ¹ H-NMR (D₂ O): δ0.95(t,6H);1.62-1.75(m,4H); 2.05-2.16(m,4H); 3.02(t,4H); 3.11-3.18(m,8H);3.76(d,4H); 6.04-6.06(m,2H).

The starting materials are prepared as follows:

a) (E)-1,14-Di-propyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene and(E)-1-propyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

0.22 g (5.5 mmol) of sodium hydride dispersion (approx. 60%) is added,with stirring, to a solution of 1.5 g (2.5 mmol) of(E)-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene in 20 ml ofDMF. The mixture is stirred for 1 h at room temperature and is cooled to10° C.; 0.5 ml (5.5 mmol) of propyl bromide is added, and the reactionmixture is stirred for a further 20 h at room temperature and is thenconcentrated by evaporation in vacuo. The residue is partitioned betweenethyl acetate and water, and the ethyl acetate phase is washed withwater and brine, is dried over sodium sulfate and is concentrated byevaporation in vacuo. The resinous residue is purified by flashchromatography over silica gel having a particle size of 0.04-0.063 mm,using ethyl acetate-hexane (1:3). The product-containing fractions areconcentrated by evaporation, yielding the 1,14-dipropyl title compound,R_(f) value=0.57 (silica gel/ethyl acetate:hexane (1:1)) and the1-propyl title compound, R_(f) value=0.46 (silica gel/ethylacetate:hexane (1:1)) in the form of colourless resins.

b )(E)-1,14-Di-propyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-enecan also be obtained as follows:

0.4 g (10 mmol) of sodium hydride dispersion (approx. 60%) is added,with stirring, to a solution of 1.5 g (2.5 mmol) of(E)-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene in 20 ml ofDMF. The mixture is stirred for 15 min. at room temperature, and then0.91 ml (10 mmol) of propyl bromide is added to the reaction mixture andstirring is continued for 41 h at room temperature. Working upanalogously to 1a) yields the title compound in the form of a colourlessresin, R_(f) value=0.57 (silica gel/ethyl acetate:hexane (1:1)).

c) (E)-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

A solution of 25.86 g (105 mmol) of2-(BOC-oxyimino)-2-phenylacetonitrile (Fluka, Switzerland) in 70 ml ofTHF is added dropwise over a period of 2 h, with stirring, to asolution, cooled to 5° C., of 5.01 g (25 mmol) of(E)-1,5,10,14-tetraazatetradec-7-ene in 50 ml of THF. The reactionmixture is stirred for a further 16 h at room temperature and is thenconcentrated by evaporation in vacuo. The residue is purified by flashchromatography, over silica gel having an average particle size of0.04-0.063 mm, using ethyl acetate-hexane (1:3). Concentration of theproduct-containing fractions by evaporation yields the resinous titlecompound, R_(f) value=0.33 (silica gel/ethyl acetate:hexane (1:1)).

EXAMPLE 2 (E)-1,14-Di-allyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

The title compound is obtained analogously to Example 1 starting from0.81 g (1.19 mmol) of(E)-1,14-di-allyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 12 ml of 3N methanolic hydrochloric acid; m.p. >260° C. ¹ H-NMR (D₂O): δ2.02-2.17(m,4H); 3.12-3.19(m,8H); 3.69(d,4H); 3.77(d,4H);5.48-5.55(m,4H); 5.83-5.97(m,2H); 6.05-6.07(m,2H).

The starting material is prepared as follows:

a) (E)-1,14-Di-ally-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand (E)-1-allyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

To a solution of 1.8 g (3 mmol) of(E)-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene in 25 ml of DMFthere are added, with stirring, 0.48 g (12 mmol) of sodium hydridedispersion (approx. 60%) and, after 15 min., 1.02 ml (12 mmol) of allylbromide. The reaction mixture is stirred for 15 h at room temperatureand for 20 h at 40° C.; a further 0.24 g (6 mmol) of sodium hydridedispersion (approx. 60%) and 0.51 ml (6 mmol) of allyl bromide are addedat 25° C. and the reaction mixture is stirred for a further 20 h at 40°C. and is then concentrated by evaporation in vacuo. Working upanalogously to Example 1a) yields the 1,14-di-allyl title compound,R_(f) value=0.57 (silica gel/ethyl acetate:hexane (1:1)), and the1-allyl title compound, R_(f) value=0.46 (silica gel/ethylacetate:hexane (1:1)), in the form of viscous oils.

EXAMPLE 3 (E)-1,14-Di-butyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

The title compound is obtained analogously to Example 1 starting from1.56 g (2.19 mmol) of(E)-1,14-di-butyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 24 ml of 3N methanolic hydrochloric acid; m.p. >260° C. ¹ H-NMR (D₂O): δ0.91(t,6H); 1.32-1.44(m,4H); 1.60-1.70(m,4H); 2.05-2.16(m,4H);3.03-3.18(m,12H) 3.77(d,4H); 6.04-6.07(m,2H).

The starting material is prepared as follows:

a)(E)-1,14-Di-butyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand (E)-1-butyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

To a solution of 1.8 g (3 mmol) of(E)-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene in 25 ml of DMFthere are added, with stirring, 0.48 g (12 mmol) of sodium hydridedispersion (approx. 60%) and, after 5 min., 1.29 ml (12 mmol) of butylbromide. The reaction mixture is stirred for 60 h at room temperature; afurther 0.24 g (6 mmol) of sodium hydride dispersion (approx. 60%) and0.645 ml (6 mmol) of butyl bromide are added, and the mixture is stirredfor a further 24 h at room temperature and is then concentrated byevaporation in vacuo. Working up analogously to Example 1a) yields the1,4-di-butyl title compound, R_(f) value=0.62 (silica gel/ethylacetate:hexane (1:1)), and the 1-butyl title compound, R_(f) value=0.49(silica gel/ethyl acetate:hexane (1:1)), in the form of colourless oils.

EXAMPLE 4 (E)-1,14-Di-propargyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

0.97 g (1.43 mmol) of(E)-1,14-di-propargyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 15 ml of 3N methanolic hydrochloric acid are reacted analogously toExample 1. Recrystallisation of the crude product from methanol/wateryields the title compound, m.p. >260° C. ¹ H-NMR (D₂ O):δ2.07-2.19(m,4H); 3.01(t,2H); 3.15-3.30(m,8H); 3.77(d,4H); 3.96(d,4H);6.05-6.07(m,2H).

The starting material is prepared as follows:

a)(E)-1,14-Di-propargyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand (E)-1-propargyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

To a solution of 1.8 g (3 mmol) of(E)-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene in 25 ml of DMFthere are added, with stirring, 0.48 g (12 mmol) of sodium hydridedispersion (approx. 60%) and, after 10 min., 0.9 ml (12 mmol) ofpropargyl bromide. The reaction mixture is stirred for 15 h at roomtemperature; a further 0.24 g (6 mmol) of sodium hydride dispersion(approx. 60%) and 0.45 ml (6 mmol) of propargyl bromide are added, andthe mixture is stirred for a further 15 h at room temperature and isthen concentrated by evaporation in vacuo. Working up analogously toExample 1a) yields the resinous 1,4-di-propargyl title compound, R_(f)value=0.55 (silica gel/ethyl acetate:hexane (1:1)), and the resinous1-propargyl title compound, R_(f) value=0.44 (silica gel/ethylacetate:hexane (1:1)).

EXAMPLE 5 (E)-1,14-Di-ethyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

1.97 g (3 mmol) of(E)-1,14-di-ethyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 30 ml of 3N methanolic hydrochloric acid are reacted analogously toExample 1. The addition of 50 ml of diethyl ether to the reactionmixture and subsequent filtration yield the title compound, m.p. >260°C. ¹ H-NMR (D₂ O): δ1.28(t,6H); 2.04-2.15(m,4H); 3.08-3.19(m,12H);3.76(d,4H); 6.04-6.07(m,2H).

The starting material is prepared as follows:

a)(E)-1,14-Di-ethyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand (E)-1-ethyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

0.62 g (15.5 mmol) of sodium hydride dispersion (approx. 60%) is added,with stirring, to a solution of 4.22 g (7.02 mmol) of(E)-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene in 50 ml ofDMF. The mixture is stirred for 15 min. at room temperature, 0.576 ml(7.72 mmol) of ethyl bromide are added, and the reaction mixture isstirred for a further 15 h at room temperature and is then concentratedby evaporation in vacuo. The residue is partitioned between ethylacetate and water, and the ethyl acetate phase is washed with water andbrine, is dried over sodium sulfate and is concentrated by evaporationin vacuo. The oily residue is purified by means of flash chromatographyover silica gel having a particle size of 0.04-0.063 ram, using ethylacetate-hexane mixtures (1:4 and 1:3). The product-containing fractionsare concentrated by evaporation, yielding the 1,14-di-ethyl titlecompound, R_(f) value=0.50 (silica gel/ethyl acetate:hexane (1:1)), andthe 1-ethyl title compound, R_(f) value=0.42 (silica gel/ethylacetate:hexane (1:1)), in the form of oils.

EXAMPLE 6 (E)-1-Ally-14-ethyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

A mixture of 0.28 g (0.418 mmol) of(E)-1-allyl-14-ethyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 6 ml of 3N methanolic hydrochloric acid is stirred for 6 h at roomtemperature, and then 10 ml of diethyl ether are added. Working upanalogously to Example 1 yields the title compound, m.p. >260° C. ¹H-NMR (D₂ O): δ1.28(t,3H); 2.05-2.17(m,4H); 3.07-3.19(m,10H);3.69(d,2H); 3.77(d,4H); 5.49-5.55(m,2H); 5.84-5.98(m,1H);6.04-6.07(m,2H).

The starting material is prepared as follows:

a)(E)-1-Allyl-14-ethyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

To a solution of 0.272 g (0.424 mmol) of(E)-1-allyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene (seeExample 2a) in 4 ml of DMF there are added, with stirring, 0.034 g (0.85mmol) of sodium hydride dispersion (approx. 60%) and, after 5 min.,0.069 ml (0.85 mmol) of ethyl iodide. The reaction mixture is stirredfor 15 h at room temperature; a further 0.017 g (0.425 mmol) of sodiumhydride dispersion (approx. 60%) and 0.034 ml (0.421 mmol) of ethyliodide are added, and the mixture is stirred for a further 96 h at 20°C. and is then concentrated by evaporation in vacuo. Working upanalogously to Example 1a), but using ethyl acetate-hexane (1:2) in theflash chromatography, yields the title compound, R_(f) value=0.53(silica gel/ethyl acetate:hexane (1:1)).

EXAMPLE 7 (E)-1-Ethyl-14-methyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

A mixture of 0.32 g (0.498 mmol) of(E)-1-ethyl-14-methyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 6 ml of 3N methanolic hydrochloric acid is stirred for 15 h at roomtemperature, and then 10 ml of diethyl ether are added. Working upanalogously to Example 1 yields the title compound, m.p. >260° C. ¹H-NMR (D₂ O): δ1.28(t,3H); 2.03-2.17(m,4H); 2.74(s,3H);3.07-3.19(m,10H); 3.77(d,4H); 6.04-6.07(m,2H).

The starting material is prepared as follows:

a)(E)-1-Ethyl-14-methyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

To a solution of 0.46 g (0.73 mmol) of(E)-1-ethyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene (seeExample 5a) in 6 ml of DMF there are added, with stirring, 0.059 g (1.47mmol) of sodium hydride dispersion (approx. 60% ) and, after 5 min.,0.092 ml (1.47 mmol) of methyl iodide. The reaction mixture is stirredfor 15 h at room temperature; a further 0.029 g (0.725 mmol) of sodiumhydride dispersion (approx. 60%) and 0.045 ml (0.721 mmol) of methyliodide are added, and the mixture is stirred for a further 96 h at 20°C. and is then concentrated by evaporation in vacuo. Working upanalogously to Example 1a), but using ethyl acetate-hexane (1:2) in theflash chromatography, yields the title compound, R_(f) value=0.42(silica gel/ethyl acetate:hexane (1:1)).

EXAMPLE 8 (E)-1-Butyl-14-ethyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

A mixture of 0.3 g (0.438 mmol) of(E)-1-butyl-14-ethyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 6 ml of 3N methanolic hydrochloric acid is stirred for 15 h at roomtemperature, and then 10 ml of diethyl ether are added. Working upanalogously to Example 1 yields the title compound, m.p. >260° C. ¹H-NMR (D₂ O): δ0.91 (t,3H); 1.27(t, 3H); 1.32-1.45(m,2H);2.05-2.16(m,4H); 3.03-3.19(m,12H); 3.77(d,4H); 6.04-6.07(m,2H).

The starting material is prepared as follows:

a)(E)-1-Butyl-14-ethyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

0.059 g (1.47 mmol) of sodium hydride dispersion (approx. 60%) is added,with stirring, to a solution of 0.46 g (0.73 retool) of(E)-1-ethyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene (seeExample 5a) in 6 ml of DMF. The mixture is stirred for 5 min. at roomtemperature; 0.159 ml (1.47 mmol) of butyl bromide is added, and thereaction mixture is stirred for a further 15 h at 20° C. and is thenconcentrated by evaporation in vacuo. Working up analogously to Example1a) yields the title compound, R_(f) value=0.56 (silica gel/ethylacetate:hexane (1:1)).

EXAMPLE 9 (E)-1-Ethyl-14-propyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

A mixture of 0.5 g (0.745 mmol) of(E)-1-ethyl-14-propyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 10 ml of 3N methanolic hydrochloric acid is stirred for 15 h at roomtemperature, and then 20 ml of diethyl ether are added. Working upanalogously to Example 1 yields the title compound, m.p. >260° C. ¹H-NMR (D₂ O): δ0.96(t,3H); 1.28(t,3H); 1.63-1.76(m,2H); 2.05-2.16(m,4H);2.97-3.18(m,12H); 3.77(d,4H); 6.05-6.07(m,2H).

The starting material is prepared as follows:

a)(E)-1-Ethyl-14-propyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

0.05 g (1.25 mmol) of sodium hydride dispersion (approx. 60%) is added,with stirring, to a solution of 0.40 g (0.622 mmol) of(E)-1-propyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene (seeExample 1a) in 6 ml of DMF. The mixture is stirred for 5 min. at roomtemperature; 0.093 ml (1.25 mmol) of ethyl bromide is added, and thereaction mixture is stirred for a further 36 h at 20° C. and is thenconcentrated by evaporation in vacuo. Working up analogously to Example1a) yields the title compound, R_(f) value=0.55 (silica gel/ethylacetate:hexane (1:1)).

EXAMPLE 10 (E)-1,14-Di-ethyl-1,5,10,14-tetraazatetradec-7 -enetetrahydrobromide and(E)-1,14-di-ethyl1-1,5,10,14-tetraazatetradec-7-ene tetrahydrochloride

A mixture of 2.8 g (3.2 mmol) of(E)-1,14-di-ethyl-1,5,10,14-tetratosyl-1,5,10,14-tetraazatetradec-7-ene,2.8 g (29.75 mmol) of phenol and 18.2 ml of an approximately 33%solution of hydrogen bromide in glacial acetic acid is heated for 8 hunder reflux. The mixture is then cooled in an ice-bath and is filtered,and the resulting crystallisate is washed with ethanol/diethyl ether(1:1) and diethyl ether. The title compound (tetrahydrobromide) obtainedafter drying under a high vacuum at 70° C. melts at >260° C. ¹ H-NMR (D₂O): δ1.23(t,6H); 1.98-2.14(m,4H); 3.02-3.16(m,12H); 3.73(d,4H);6.01-6.04(m,2H).

For conversion into the tetrahydrochloride, 0.91 g (1.57 mmol) of(E)-1,14-di-ethyl-1,5,10,14-tetraazatetradec-7-ene tetrahydrobromide isdissolved in 2 ml of water and chromatographed over a column chargedwith ®Amberlite-IRA-400 ion exchanger (OH⁻ form; strongly basic ionexchanger based on a styrene/divinylbenzene polymer having a quaternaryammonium function), using water as eluant. The combinedproduct-containing fractions are acidified to a pH of about 2 with 2Nhydrochloric acid and are concentrated by evaporation in vacuo.Recrystallisation of the residue from ethanol/water yields the titlecompound (tetrahydrochloride), m.p. >260° C. ¹ H-NMR (D₂ O):δ1.26(t,6H); 2.05-2.14(m,4H); 3.06-3.17(m,12H); 3.76(d,4H);6.04-6.06(m,2H).

The starting materials are prepared as follows:

a)(E)-1,14-Di-ethyl-1,5,10,14-tetratosyl-1,5,10,14-tetraazatetradec-7-eneand (E)-1-ethyl-1,5,10,14-tetratosyl-1,5,10,14-tetraazatetradec-7-ene

25.4 g (0.1838 mol) of potassium carbonate (anhydrous) and 7.1 ml(0.0951 mol) of ethyl bromide are added, with stirring, to a solution of30 g (0.0367 mmol) of(E)-1,5,10,14-tetratosyl-1,5,10,14-tetraazatetradec-7-ene in 100 ml ofDMF. The reaction mixture is stirred for 22 h at 70° C., and then afurther 2.36 ml (0.0316 mol) of ethyl bromide are added and the mixtureis stirred for a further 12 h at 70° C. The reaction mixture is cooledto room temperature and is filtered, and the filtrate is concentrated byevaporation in vacuo. The residue is purified by means of flashchromatography twice over silica gel having a particle size of0.04-0.063 mm, using toluene-ethyl acetate (10:1 and 5:1 ) and ethylacetate-hexane (1:1), yielding the oily 1,14-di-ethyl title compound,R_(f) value=0.75 (silica gel/methylene chloride:methanol (50:1)), andthe oily 1-ethyl title compound, R_(f) value=0.65 (silica gel/methylenechloride:methanol (50:1)).

b) (E)-1,5,10,14-tetratosyl-1,5,10,14-tetraazatetradec-7-ene

A solution of 38.13 g (0.2 mol) of p-toluenesulfonic acid chloride in300 ml of methylene chloride is added dropwise at room temperature, withstirring and under a nitrogen atmosphere, to a solution of 10.02 g (0.05mol) of (E)-1,5,10,14-tetraazatetradec-7-ene in 100 ml (0.2 mol) of 2Nsodium hydroxide solution. The reaction mixture is stirred for a further2 h at room temperature, and then the organic phase is separated off andthe aqueous phase is extracted with methylene chloride. The organicphases are combined, washed with water, dried over sodium sulfate andconcentrated by evaporation. The residue is purified by means of flashchromatography over silica gel having a particle size of 0.04-0.063 mm,using ethyl acetate-hexane (1:1). Concentration of theproduct-containing fractions by evaporation yields the title compound inthe form of an amorphous residue, R_(f) value=0.51 (silica gel/methylenechloride:methanol (50:1)).

EXAMPLE 11 (E)-1,14-Di-ethyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

A mixture of 0.18 g (0.642 mmol) of(E)-1,4-bis(3-ethyl-hexahydropyrimidin-1-yl)but-2-ene, 2.5 ml ofmethanol and 2.56 ml (5.12 mmol) of 2N hydrochloric acid is heated for72 h under reflux. The reaction mixture is then cooled to roomtemperature, and 10 ml of methanol are added, whereupon the titlecompound precipitates in crystalline form. The crystallisate, which iswashed with methanol and diethyl ether and dried under a high vacuum at120° C., melts at >260° C. ¹ H-NMR (D₂ O): δ1.27(t,6H); 2.04-2.15(m,4H);3.07-3.18(m,12H); 3.77(d,4H); 6.04-6.06(m,2H).

The starting material is prepared as follows:

a) (E)-1,4-Bis(3-ethyl-hexahydropyrimidin-1-yl)but-2-ene

A mixture of 1.07 g (9.37 mmol) of 1-ethyl-hexahydro-pyrimidine (Chem.Pharm. Bull. 28, 3310 (1980)), 0.5 g (2.34 mmol) oftrans-1,4-dibromo-2-butene and 10 ml of acetonitrile is heated for 16 hat 80° C., with stirring and under a nitrogen atmosphere, and is thenconcentrated by evaporation in vacuo. The residue is partitioned between2N sodium hydroxide solution and methylene chloride, and the methylenechloride phase is washed with brine, dried over sodium sulfate andconcentrated by evaporation in vacuo. The residue is purified by meansof flash chromatography over silica gel having a particle size of0.04-0.063 mm, using methylene chloride-methanol (9:1) and methylenechloridemethanol-concentrated ammonia (90:10:0.5), yielding the titlecompound in the form of an oil, R_(f) value=0.63 (silica gel/methylenechloride:methanol:concentrated ammonia (40:10:1)).

EXAMPLE 12 (E)-1-Allyl-14-propargyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

A mixture of 0.22 g (0.324 mmol) of(E)-1-allyl-14-propargyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 3.3 ml of 3N methanolic hydrochloric acid is reacted analogously toExample 8. The resulting title compound melts at >260° C. ¹ H-NMR (D₂O): δ2.07-2.19(m,4H); 3.02(t,1H); 3.08-3.34(m,8H); 3.69(d,2H);3.77(d,4H); 3.96(d,2H); 5.48-5.55(m,2H); 5.84-5.97(m,1H);6.04-6.07(m,2H).

The starting material is prepared as follows:

a)(E)-1-Allyl-14-propargyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

To a solution of 0.6 g (0.936 mmol) of (E)-1-allyl-1,5, 10,14-tetra-BOC- 1,5,10,14-tetraazatetradec-7-ene (see Example 2a) in 8 mlof DMF there are added, with stirring, 0.075 g (1.875 mmol) of sodiumhydride dispersion (approx. 60%) and, after 5 min., 0.141 ml (1.872mmol) of propargyl bromide. The reaction mixture is stirred for 15 h atroom temperature; a further 0.0375 g (0.937 mmol) of sodium hydridedispersion (approx. 60%) and 0.071 ml (0.94 mmol) of propargyl bromideare added, and the mixture is stirred for a further 96 h at 20° C. andis then concentrated by evaporation in vacuo. Working up analogously toExample 1a) yields the title compound, R_(f) value=0.44 (silicagel/ethyl acetate:hexane (2:3)).

EXAMPLE 13 (E)-1-Allyl-14-propyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

A mixture of 0.55 g (0.805 mmol) of(E)-1-allyl-14-propyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 8.2 ml of 3N methanolic hydrochloric acid is reacted analogously toExample 8. The resulting title compound melts at >260° C. ¹ H-NMR (D₂O): δ0.97(t,3H); 1.63-1.76(m,2H); 2.05-2.16(m,4H); 3.02(t,2H);3.11-3.19(m,8H); 3.69(d,2H); 3.77(d,4H); 5.48-5.55(m,2H);5.83-5.98(m,1H); 6.04-6.07(m,2H).

The starting material is prepared as follows:

a)(E)-1-Allyl-14-propyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

To a solution of 0.7 g (1.092 mmol) of(E)-1-allyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene (seeExample 2a) in 9.4 ml of DMF there are added, with stirring, 0.087 g(2.175 mmol) of sodium hydride dispersion (approx. 60%) and, after 5min., 0.212 ml (2.176 mmol) of propyl iodide. The reaction mixture isstirred for 15 h at room temperature; a further 0.043 g (1.075 mmol) ofsodium hydride dispersion (approx. 60%) and 0.106 ml (1.088 mmol) ofpropyl iodide are added, and the mixture is stirred for a further 96 hat 20° C. and is then concentrated by evaporation in vacuo. Working upanalogously to Example 1a) yields the oily title compound, R_(f)value=0.46 (silica gel/ethyl acetate:hexane (2:3)).

EXAMPLE 4 (E)-1-Allyl-14-methyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

A mixture of 0.6 g (0,916 mmol) of(E)-1-allyl-14-methyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 9 ml of 3N methanolic hydrochloric acid is reacted analogously toExample 8. The resulting title compound melts at >260° C. ¹ H-NMR (D₂O): δ2.07-2.17(m,4H); 2.73(s,3H); 3.12-3.19(m,8H); 3.69(d,2H);3.77(d,4H); 5.48-5.55(m,2H); 5.83-5.98(m, 1H); 6.04-6.07 (m,2H).

The starting material is prepared as follows:

a)(E)-1-Allyl-14-methyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

To a solution of 0.6 g (0.936 mmol) of(E)-1-allyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene (seeExample 2a) in 8 ml of DMF there are added, with stirring, 0.075 g(1.875 mmol) of sodium hydride dispersion (approx. 60%) and, after 5min., 0.117 ml (1.875 mmol) of methyl iodide. The reaction mixture isstirred for 15 h at room temperature; a further 0.075 g (1.875 mmol) ofsodium hydride dispersion (approx. 60% ) and 0.117 ml (1.875 mmol) ofmethyl iodide are added, and the mixture is stirred for a further 96 hat 20° C. and is then concentrated by evaporation in vacuo. Working upanalogously to Example 1a, but using ethyl acetate-hexane (1:2) in theflash chromatography, yields the title compound, R_(f) value=0.36(silica gel/ethyl acetate:hexane (2:3)).

EXAMPLE 15 (E)-1-Isopropyl-14-propyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

A mixture of 0.24 g (0.35 mmol) of(E)-1-isopropyl-14-propyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 4 ml of 3N methanolic hydrochloric acid is stirred for 4 h at roomtemperature. Then the reaction mixture is diluted with 6 ml of diethylether and is worked up analogously to Example 1. The resulting titlecompound melts at >260° C. ¹ H-NMR (D₂ O): δ0.97(t,3H); 1.32(d,6H);1.63-1.76(m,2H); 2.04-2.17(m,4H); 3.02(t,2H); 3.12-3.19(m,8H);3.37-3.49(m,1H); 3.78(d,4H); 6.05-6.07(m,2H).

The starting materials are prepared as follows:

a)(E)-1-Isopropyl-14-propyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene0.042 g (1.05 mmol) of sodium hydride dispersion (approx. 60%) is added,with stirring, to a solution of 0.338 g (0.526 mmol) of(E)-1-isopropyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene in5 ml of DMF. The mixture is stirred for 5 min. at room temperature;0.103 ml (1.05 mmol) of propyl iodide is added, and the reaction mixtureis stirred for a further 16 h at 20° C. and is then concentrated byevaporation in vacuo. Working up analogously to Example 1a), using ethylacetate-hexane (1:3) and ethyl acetate-hexane (1:2) in the flashchromatography, yields the title compound, R_(f) value=0.51 (silicagel/ethyl acetate:hexane (2:3)).

b) (E)-1-Isopropyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

To a solution of 1.5 g (2.5 mmol) of(E)-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec7-ene in 20 ml of DMFthere are added, with stirring, 0.4 g (10 mmol) of sodium hydridedispersion (approx. 60%) and, after 15 min., 1 ml (10 mmol) of isopropyliodide. The reaction mixture is stirred for 16 h at room temperature; afurther 0.4 g (10 mmol) of sodium hydride dispersion (approx. 60%) and 1ml (10 mmol) of isopropyl iodide are added, and the mixture is stirredfor a further 24 h at 20° C. and for 24 h at 50° C. and is thenconcentrated by evaporation in vacuo. Working up analogously to Example1a), using ethyl acetate-hexane (1:3) and ethyl acetate-hexane (1:2) inthe flash chromatography, yields the title compound, R_(f) value=0.37(silica gel/ethyl acetate:hexane (2:3)).

EXAMPLE 16(E)-1,14-Di-cyclopropylmethyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

A mixture of 1.5 g (2.116 mmol) of(E)-1,14-di-cyclopropylmethyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-eneand 23 ml of 3N methanolic hydrochloric acid is stirred for 5 h at roomtemperature, and then 20 ml of diethyl ether are added. Working upanalogously to Example 1 yields the title compound, m.p. >260° C. ¹H-NMR (D₂ O): δ0.33-0.38(m,4H); 0.65-0.71 (m,4H); 1.00-1.14(m,2H);2.05-2.16(m,4H); 2.96(d,4H); 3.13-3.19(m,8H); 3.77(d,4H);6.04-6.07(m,2H).

The starting material is prepared as follows:

a)(E)-1,14-Di-cyclopropylmethyl-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene

0.4 g (10 mmol) of sodium hydride dispersion (approx. 60%) is added,with stirring, to a solution of 1.5 g (2.5 mmol) of(E)-1,5,10,14-tetra-BOC-1,5,10,14-tetraazatetradec-7-ene in 21.5 ml ofDMF. The mixture is stirred for 15 min. at room temperature; then 1.06ml (10 mmol) of bromomethyl-cyclopropane (approx. 90% ) are added to thereaction mixture and stirring is continued for 90 h at room temperature.Working up analogously to Example 1a) yields the title compound in theform of a colourless resin, R_(f) value=0.47 (silica gel/ethylacetate:hexane (2:3)).

EXAMPLE 17 (E)-1,14-Di-propyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

A mixture of 0.265 g (0.859 mmol) of(E)-1,4-bis(3-propylhexahydro-pyrimidin-1-yl)but-2-ene, 3 ml of methanoland 3.4 ml (6.8 mmol) of 2N hydrochloric acid is heated for 19 h underreflux and is then concentrated by evaporation in vacuo.Recrystallisation of the residue from methanol/water yields the titlecompound, m.p. >260° C. ¹ H-NMR (D₂ O): δ0.96(t,6H); 1.62-1.76(m,4H);2.05-2.16(m,4H); 3.02(t,4H); 3.11-3.18(m,8H); 3.77(d,4H);6.03-6.06(m,2H).

The starting materials are prepared as follows:

a) (E)-1,4-Bis(3-propylhexahydro-pyrimidin-1-yl)but-2-ene

A mixture of 0.68 g (17.9 mmol) of lithium aluminium hydride, 10 ml ofdioxane and 1 g (2.97 mmol) of(E)-1,4-bis(3-propionylhexahydro-pyrimidin-1-yl)but-2-ene is stirred for15 h at 100° C., with stirring and under a nitrogen atmosphere. Aftercooling to 5° C. in an ice-bath, there are added dropwise to thereaction mixture, in succession, a mixture of 0.53 ml of water and 2 mlof tetrahydrofuran, 0.53 ml of 1N sodium hydroxide solution and 1.1 mlof water, care being taken to ensure that the temperature does notexceed 15° C. The mixture is stirred for a further 1 h at roomtemperature and is filtered, the filtration residue is washed withtetrahydrofuran, and the filtrate is then concentrated by evaporation invacuo. The oily residue is purified by means of flash chromatographyover silica gel having a particle size of 0.04-0.063 mm, using methylenechloride-methanol (10:1) and methylene chloride-methanol-concentratedammonia (100:10:0.5). Concentration of the product-containing fractionsby evaporation yields the title compound in the form of an oil, R_(f)value=0.56 (silica gel/methylene chloride:methanol:concentrated ammonia(150:50:1)).

b) (E)-1,4-Bis(3-propionylhexahydro-pyrimidin-1-yl)but-2-ene

5 ml (38.8 mmol) of propionic acid anhydride are added at 0° C., withstirring, to a mixture of 1.77 g (7.89 mmol) of(E)-1,4-bis(hexahydro-pyrimidin-1-yl)but-2-ene and 10 ml of pyridine.The reaction mixture is stirred for a further 1 h at room temperatureand is then concentrated by evaporation in vacuo. The oily residue ispartitioned between 15% sodium hydroxide solution and ethyl acetate. Theorganic phase is washed with brine and dried over sodium sulfate and isthen concentrated by evaporation in vacuo, yielding the title compoundin the form of an oil, R_(f) value=0.77 (silica gel/methylenechloride:methanol:concentrated ammonia (40:10:1)).

c) (E)-1,4-Bis(hexahydro-pyrimidin-1-yl)but-2-ene

A mixture of 7.6 ml (99.8 mmol) of formaldehyde (approx. 36.5% aqueoussolution) and 10 ml of water is added dropwise over a period of 1 h,with stirring, to a solution, cooled to 0° C., of 10 g (49.9 mmol) of(E)-1,5,10,14-tetraazatetradec-7-ene in 25 ml of water. The reactionmixture is stirred for a further 2 h at 0° C. and a further 15 h at roomtemperature and is then concentrated by evaporation in vacuo. Theresidue is purified by means of flash chromatography over silica gelhaving a particle size of 0.04-0.063 mm, using methylenechloride-methanol-concentrated ammonia (100:10:0.5) and methylenechloride-methanol-concentrated ammonia (50:10:1). The product-containingfractions are concentrated by evaporation and the residue is sublimedunder a high vacuum at 85° C. The resulting title compound melts at125°-127° C.

EXAMPLE 18 (E)-1,14-Di-propyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

0.25 g (0.0008 mol) of(E)-1,14-di-propionyl-1,5,10,14-tetraazatetradec-7-ene is added, withstirring and while cooling in an ice-bath, to a solution of 2.4 g(approx. 0.00831 mol) of sodium dihydro-bis(2-methoxyethoxy)aluminate(approx. 70% solution in toluene; Fluka, Buchs, Switzerland) in 10 ml ofTHF. The reaction mixture is heated for 2 h under reflux and is thencooled in an ice-bath, and 2.4 ml of 30% sodium hydroxide solution, 10ml of THF and 3 g of anhydrous magnesium sulfate are added insuccession. The mixture is filtered, the filtration residue is washedwith THF, and the filtrate is concentrated by evaporation in vacuo. Theoily residue is dissolved in 15 ml of ethanol, and 1 ml of 4Nhydrochloric acid is added to the solution. The crystallisate that formsis filtered off and then recrystallised twice more from methanol/water.The resulting title compound melts at >260° C. ¹ H-NMR (D₂ O):δ0.96(t,6H); 1.63-1.76(m,4H); 2.05-2.16(m,4H); 3.02(t,4H);3.11-3.18(m,8H); 3.77(d,4H); 6.04-6.07(m,2H).

The starting material is prepared as follows:

a) (E)-1,14-Di-propionyl-1,5,10,14-tetraazatetradec-7-ene

A mixture of 1.84 g (0.005468mmol) of(E)-1,4-bis(3-propionylhexahydro-pyrimidin-1-yl)but-2-ene (Example 17b),44 ml of methanol, 2.74 ml (0.03405 mol) of pyridine and 4.11 g (0.03949mol) of malonic acid is heated for 2 h under reflux and is thenconcentrated by evaporation in vacuo. 10 ml of 30% sodium hydroxidesolution are added to the oily residue, the mixture is extractedthoroughly with ethyl acetate, the combined ethyl acetate extracts aredried over anhydrous sodium sulfate, and the resulting product isconcentrated by evaporation in vacuo to a volume of about 25 ml. Thecrystalline precipitate that forms is filtered off and, for purificationpurposes, is recrystallised from acetonitrile. The resulting titlecompound melts at 82°-84° C.

EXAMPLE 19 (E)-1,14-Di-ethyl-1,5,10,14-tetraazatetradec-7-enetetrahydrochloride

0.5 g (0.624 mmol) of(E)-1,14-di-ethyl-1,5,10,14-tetra-diethoxyphosphoryl-1,5,10,14-tetraazatetradec-7-eneis dissolved in 5 ml of THF; then hydrogen chloride is passed into thesolution for a period of about 45 min. at 0°-5° C. and the reactionmixture is stirred for 20 h at room temperature. The addition of 15 mlof diethyl ether to the reaction mixture, filtration, washing thecrystallisate with diethyl ether and drying under a high vacuum yieldthe title compound, m.p. >260° C. ¹ H-NMR (D₂ O): δ1.28(t,6H);2.05-2.16(m,4H); 3.08-3.19(m,12H); 3.77(d,4H); 6.05-6.07(m,2H).

The starting materials are prepared as follows:

a)(E)-1,14-Di-ethyl-1,5,10,14-tetra-diethoxyphosphoryl-1,5,10,14-tetraazatetradec-7-ene

0.16 g (4 mmol) of sodium hydride dispersion (approx. 60%) is added,with stirring, to a solution of 0.745 g (1 mmol) of(E)-1,5,10,14-tetra-diethoxyphosphoryl-1,5,10,14-tetraazatetradec-7-enein 5 ml of DMF. The mixture is stirred for 10 min. at room temperature,0.3 ml (4 mmol) of ethyl bromide are added, and the reaction mixture isstirred for a further 2 h at 20° C. and is then concentrated byevaporation in vacuo. The residue is partitioned between ethyl acetateand ice-cold water, and the ethyl acetate phase is washed with water andbrine, is dried over sodium sulfate and is concentrated by evaporationin vacuo. The oily residue is purified by means of flash chromatographyover silica gel having a particle size of 0.04-0.063 mm, using methylenechloride-methanol mixtures (49:1 and 9:1). Concentration of theproduct-containing fractions by evaporation yields the title compound inthe form of a colourless oil, R_(f) value=0.34 (silica gel, eluantmethylene chloride:methanol (9:1)).

b)(E)-1,5,10,14-Tetra-diethoxyphosphoryl-1,5,10,14-tetraazatetradec-7-ene

A solution of 12.4 g (89.8 mmol) of diethyl phosphite in 18 ml of carbontetrachloride is added dropwise at 10°-15° C., with stirring, to amixture of 3 g (14.97 mmol) of (E)-1,5,10,14-tetraazatetradec-7-ene, 18g (179.8 mmol)of potassium hydrogen carbonate, 24.85 g (179.8 mmol) ofpotassium carbonate, 1.45 g (4.5 mmol) of tetrabutylammonium bromide and70 ml of methylene chloride. The reaction mixture is stirred for afurther 90 h at room temperature and is filtered, the filtrate is washedwith water, and the organic phase is dried over sodium sulfate andconcentrated by evaporation in vacuo. The oily residue is purified bymeans of flash chromatography over silica gel having a particle size of0.04-0.063 mm, using methylene chloride-methanol (49:1) and methylenechloridemethanol (9:1). Concentration of the product-containingfractions by evaporation yields the title compound in the form of anoil, R_(f) value=0.28 (silica gel, eluant methylene chloride:methanol(9:1)).

EXAMPLE 20

(E)-1,14-Di-allyl-1,5,10,14-tetraazatetradec-7-ene tetrahydrochloride

For conversion into the title compound, 37.45 g of crude(E)-1,14-di-allyl-1,5,10,14-tetra(diethoxyphosphoryl)-1,5,10,14-tetraazatetradec-7-eneare dissolved in 170 ml of 7THF, and hydrogen chloride is passed intothe solution at 5° C. until saturated. The reaction mixture is stirredfor 15 h at room temperature, 250 ml of diethyl ether are added to theresulting suspension, the mixture is filtered and the filtration residueis washed with a small amount of ethanol and diethyl ether.Recrystallisation from ethanol/water (with treatment with activatedcarbon) and drying the crystallisate under a high vacuum at 120° C.yield the title compound, m.p. >260° C. ¹ H-NMR (D₂ O):δ2.06-2.17(m,4H); 3.13-3.19(m,8H); 3.69(d,4H); 3.78(d,4H);5.49-5.55(m,4H); 5.84-5.98(m,2H); 6.05-6.07(m,2H).

The starting materials are prepared as follows:

a) (E)-1,5,10,14-Tetra(diethoxyphosphoryl)-1,5,10,14-tetraazatetradec-7-ene

A solution of 43.5 g (0.315 mol) of diethyl phosphite in 90 ml of carbontetrachloride is added dropwise at 10°-15° C. over a period of 45 min.,with stirring, to a mixture of 15.02 g (0.075 mol) of(E)-1,5,10,14-tetraazatetradec-7-ene, 63.08 g (0.63 mmol) of potassiumhydrogen carbonate, 87.07 g (0.63 mol) of potassium carbonate, 5.093 g(0.0158 mol) of tetrabutylammonium bromide and 350 ml of methylenechloride. The reaction mixture is stirred for a further 87 h at roomtemperature and is then allowed to stand for 63 h at 20° C. and isfiltered. The filtrate is washed with water and the organic phase isdried over sodium sulfate and is concentrated by evaporation in vacuo.The evaporation residue that is obtained is crude(E)-1,5,10,14-tetra(diethoxyphosphoryl)-1,5,10,14-tetraazatetradec-7-enein the form of a yellow oil, R_(f) value=0.28 (silica gel/methylenechloride:methanol (9:1)), which is used further directly.

b)(E)-1,14-Di-allyl-1,5,10,14-tetra(diethoxyphosphoryl)-1,5,10,14-tetraazatetradec-7-ene

51.3 g of crude(E)-1,5,10,14-tetra(diethoxyphosphoryl)-1,5,10,14-tetraazatetradec-7-eneare dissolved in 100 ml of DMF, and 8.27 g (0.2067 mol) of sodiumhydride dispersion (approx. 60%) are added in portions to the solution,with stirring. After stirring for 15 min. at 20° C., 17.49 ml (0.2067mol) of allyl bromide are added dropwise, and the reaction mixture isstirred for a further 15 h at room temperature and is then concentratedby evaporation in vacuo. The evaporation residue is partitioned betweenethyl acetate and ice-cold water, and the organic phase is washed insuccession with 20% citric acid, 2N sodium carbonate solution and brine,is dried over sodium sulfate and is concentrated by evaporation invacuo. There is obtained as residue crude(E)-1,14-di-allyl-1,5,10,14-tetra(diethoxyphosphoryl)-1,5,10,14-tetraazatetradec-7-enein the form of a yellow oil, R_(f) value=0.38 (silica gel/methylenechloride:methanol (9:1)).

EXAMPLE 21 Capsules

Capsules each comprising 1 g of active ingredient, for example one ofthe acid addition salts of Examples 1 to 20, can be prepared as follows:

    ______________________________________                                        Composition (for 1250 capsules):                                              ______________________________________                                        active ingredient     1250 g                                                  talc                  180 g                                                   wheat starch          120 g                                                   magnesium stearate     80 g                                                   lactose                20 g                                                   ______________________________________                                    

The powdered substances are pressed through a sieve having a mesh sizeof 0.6 mm and mixed 1.32 g portions of the mixture are introduced intogelatin capsules by means of a capsule-filling machine.

EXAMPLE 22 Levels of Putrescine, spermidine and spermine in mouseascites L 1210 leukemia cells:

Using the method analoguous to Porter et al. as described in detailabove, the following levels of putrescine (PU), spermidine (SPD) andspermine (SPM) (given in % of the respective control without addition ofa compound of formula I) are obtained when the title compound isadministered in the respective concentration (μM) given in the table (48h incubation):

    ______________________________________                                                                         concentration                                Example % PU    % SPD     % SPM  (μM) of test cpd.:                        ______________________________________                                        1, 17,  9.4     12        67     10                                           18                                                                            2, 20   16      3.1       36     10                                           5, 10,  4.1     2.1       30      5                                           11, 19                                                                        6       4.1     2.1       12     10                                           8       4.8     0.2       4.0    10                                           9       6.9     2.0       19     10                                           13      0       1.3       25     10                                           16      12      24        62     10                                           ______________________________________                                    

EXAMPLE 23 Inhibition of growth of human T24 bladder carcinoma cells:

Using the method for inhibition of growth of human T24 bladder carcinomacells om Eagle's minimal essential medium (see above), the followingtest results are obtained (given as IC₅₀ =concentration of the activeingredient at which the number of cells per well at the end of theincubation period is only 50% of the number of cells in the controlcultures):

    ______________________________________                                        Compound of Example No.                                                                           IC.sub.50 (μM)                                         ______________________________________                                        1, 17, 18           0.64                                                      2, 20               0.60                                                      3                   8.2                                                       6                   3                                                         8                   0.55                                                      9                   1.1                                                       13                  2.14                                                      16                  0.92                                                      ______________________________________                                    

What is claimed is:
 1. A compound of formula I, ##STR9## wherein R₁ andR₂, each independently of the other, are selected from C₂ -C₄ alkyl, C₃-C₄ alkenyl wherein the double bond does not originate from the carbonatom that is bonded to a nitrogen bonding R₁ or R₂, and from C₃ -C₄cycloalkylmethyl, or a salt thereof.
 2. A compound according to claim 1with the proviso that R₁ and R₂ together have not more than 6 carbonatoms.
 3. A compound according to claim 1 wherein R₁ and R₂, eachindependently of the other, are selected from ethyl, n-propyl, n-butyland allyl, or a salt thereof.
 4. A compound according to claim 3,wherein R₁ and R₂ together have 4, 5 or 6 carbon atoms, or a saltthereof.
 5. A compound according to claim 1, wherein R₁ is ethyl and R₂is selected from ethyl, n-propyl, n-butyl and allyl, or a salt thereof.6. A pharmaceutical composition suitable for administration to awarm-blooded animal for the treatment of proliferative disorders thatare responsive to a reduction in the intracellular polyamineconcentration of natural polyamines, comprising a compound of formula Iaccording to claim 1, or a pharmaceutically acceptable salt of such acompound having at least one salt-forming group that is effective in thetreatment of the mentioned disorder, together with a pharmaceuticallyacceptable carrier.
 7. A method of treating a warm-blooded animalsuffering from bladder carcinoma that is responsive to a reduction inthe intracellular concentration of polyamines, which method comprisesadministering to the warm-blooded animal requiring such treatment acompound of formula I according to claim 1, or a pharmaceuticallyacceptable salt thereof, in a dose that is effective in reducing theintracellular concentration of polyamines.
 8. A process for thepreparation of a compound of formula I according to claim 1, whichprocess comprisesa) nucleophilically substituting an amino compound offormula II ##STR10## wherein R₂ is as defined for compounds of formula Iand any functional groups that are not to take part in the reaction are,if necessary, in protected form, with a compound of formula III

    R.sub.1 --X                                                (III),

wherein R₁ is as defined for compounds of formula I and X is anucleofugal leaving group, and removing any protecting groups that arepresent,and, if desired, converting an obtainable free compound offormula I into its salt, converting an obtainable salt of a compound offormula I into the free compound or into a different salt of a compoundof formula I, and/or separating obtainable mixtures of isomers into theindividual isomers.
 9. The compound according to claim 1, said compoundbeing selected from the group consistingof(E)-1,14-Di-allyl-1,5,10,14-tetraazatetradec-7-ene;(E)-1,14-Di-butyl-1,5,10,14-tetraazatetradec-7-ene of formula I;(E)-1,14-Di-ethyl-1,5,10,14-tetraazatetradec-7-ene;(E)-1-Allyl-14-ethyl-1,5,10,14-tetraazatetradec-7-ene;(E)-1-Butyl-14-ethyl-1,5,10,14-tetraazatetradec-7-ene;(E)-1-Ethyl-14-propyl-1,5,10,14-tetraazatetradec-7-ene;(E)-1-Allyl-14-methyl-1,5,10,14-tetraazatetradec-7-ene;(E)-1-Isopropyl-14-propyl-1,5,10,14-tetraazatetradec-7-ene;(E)-1,14-Di-cyclopropylmethyl-1,5,10,14-tetraazatetradec-7-ene; and(E)-1,14-Di-propyl-1,5,10,14-tetraazatetradec-7-ene; or a salt thereof.10. The compound according to claim 1, said compound being(E)-1,14-di-propyl-1,5,10,14-tetraazatetradec-7-ene, or apharmaceutically acceptable salt thereof.
 11. The compound according toclaim 1, said compound being(E)-1-allyl-14-propyl-1,5,10,14-tetraazatetradec-7-ene, or apharmaceutically acceptable salt thereof.